Transgenics For Delayed Fruit Ripening
By
Sukanya 1385
Highly coordinated
Genetically programmed
Irreversible phenomenon
Physiological biochemical changes
Development of a soft and edible ripe fruit
What is Fruit Ripening
The molecular mechanisms controlling the ripening of fruit
Open Archive TOULOUSE Archive Ouverte
Increased respiration
Chlorophyll degradation
Biosynthesis of carotenoids anthocyanins essential
oils Flavor and aroma Components
Increased activity of cell wall-degrading enzymes
Transient increase in ethylene production
What are the changes
Major Developmental Changes during Tomato Fruit
Development and Ripening
The Plant Cell Vol 16 S170ndashS180 2004
Based on their respiratory pattern and ethylene biosynthesis
during ripening
VPrasanna et al
Classification of fruits
Pathway for ethylene biosynthesis
Rate limiting step
Critical Reviews in Food Science and Nutrition 471ndash19 2007
The expression of
ethylene biosynthesis
and ethylene
perception genes
during the transition to
climacteric in tomato
Kevany et al 2007
Bleecker and Kende 2000
Ethylene Perception and Signal Transduction
Fruit pulp or the mesocarp
parenchymatus cells
complex network of polysaccharides and proteins
The primary cell wall contains
35 pectin
25 cellulose
20 hemicellulose
10 structural hydroxyproline-rich protein
Structural components of fruits
Plant polysaccharides play a major role in
storage mobilization of energy and in
maintaining cell and tissue integrity due to
their structural and water binding capacity
Enzymes Related to Pectin Dissolution
Critical Reviews in Food Science and Nutrition 471ndash19 (2007)
Globally cultivated fleshy fruit
Worldrsquos largest vegetable crop after potato
Indian production scenario-
350000 hectares 5300000 tonsyear
Short generation time 3-4 months
Simple genetics
Numerous characterized mutants
Cross fertile wild germplasm to promote genetic studies
Routine transformation technology
Postharvest losses-5 to 25 in developed countries
-20 to 50 in developing countries
Tomato model systems for fruit development
and ripening
Natural Mutants Affected in the Ripening Phenotype
Disadvantages of existing methods of storage
Labor intensive
costly
Occupies a large floor space
Poor heat transfer may occur resulting in poor product quality
Excessive dehydration in unpacked products
Chemical changes during freezing
-enzyme-activated browning
-development of rancid oxidative flavors
Textural changes during freezing
-mushy and watery
The use of 1-methylcyclopropene (1-MCP) on fruits and
vegetables
Inhibitor of ethylene perception
Easily released as a gas when the powder is dissolved in water
Approved by the Environmental Protection Agency (EPA) in 1999
Marketed as EthylBlocreg by Floralife Inc (Walterboro SC)AgroFreshInc a
subsidiary of Rohm and Haas (Springhouse PA)
CB WatkinsBiotechnology Advances 24 (2006) 389ndash409
Transgenic approach
Delayed fruit
ripening
BLOCKING THE PERCEPTION OF ETHYLENE
BLOCKING THE EXPRESSION
OF GENES INDUCED BY ETHYLENE
BLOCKING ETHYLENE SYNTHESIS
Regulation of Ethylene Production
a Suppression of ACC synthase gene expression
ACC (1-aminocyclopropane-1-carboxylic acid) (ACS2)
conversion of S-adenosylmethionine (SAM) to ACC
the second to the last step in ethylene biosynthesis
an antisense (ldquomirror-imagerdquo) or truncated copy of the synthase gene
Oeller et al 1991
Yao et al1999
Nath et al 2006
Antisense Technology
httpagbiosafetyunleduflashantisenseswf Journal of Plant Physiology170987ndash 9952013
Null Mutation of the MdACS3 Gene
Apple cultivars homozygous or heterozygous for null allelotype
showed no or very low expression of ripening-related genes and
maintained fruit firmness
Aide Wang 2009
RNAi-mediated silencing
Chimeric RNAi-ACS construct designed to target ACS
homologs
Delayed ripening and extended shelf life for sim45 days
Aarti Gupta Ram Krishna Pal Delayed ripening and improved fruit processing quality in tomato by RNAi-mediated silencing of three
homologs of 1-aminopropane-1-carboxylate synthase gene Journal of Plant Physiology 170 (2013) 987ndash 995
Regulation of Ethylene Production
b Suppression of ACC oxidase gene expression
It catalyzes the oxidation of ACC to ethylene
The last step in the ethylene biosynthetic pathway
Down regulation through anti-sense technology
Hamilton et al 1990
Ye et al 1996
Xiong et al 2003
Ripening in papaya fruit is altered by ACC
oxidase cosuppression
Fig1Map of the construct pKYCPACOO-1 containing the ACC oxidase
fragment cloned in PKYLX80 in the sense orientation The ACC oxidase
fragment is flanked by the CaMV 35S promoter and the RUBISCO
terminator
Fig2 Ethylene production in papaya transgenic fruits
Rodolfo Loacutepez-Goacutemezet alTransgenic Res 1889ndash97 2009
c Insertion of the ACC deaminase gene
Regulation of Ethylene Production
The gene is obtained from Pseudomonas chlororaphis
(a common nonpathogenic soil bacterium)
It converts ACC to a different compound
Reduce the amount of ACC available for ethylene production
90-97 reduced ethylene production
Klee et al1991
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Highly coordinated
Genetically programmed
Irreversible phenomenon
Physiological biochemical changes
Development of a soft and edible ripe fruit
What is Fruit Ripening
The molecular mechanisms controlling the ripening of fruit
Open Archive TOULOUSE Archive Ouverte
Increased respiration
Chlorophyll degradation
Biosynthesis of carotenoids anthocyanins essential
oils Flavor and aroma Components
Increased activity of cell wall-degrading enzymes
Transient increase in ethylene production
What are the changes
Major Developmental Changes during Tomato Fruit
Development and Ripening
The Plant Cell Vol 16 S170ndashS180 2004
Based on their respiratory pattern and ethylene biosynthesis
during ripening
VPrasanna et al
Classification of fruits
Pathway for ethylene biosynthesis
Rate limiting step
Critical Reviews in Food Science and Nutrition 471ndash19 2007
The expression of
ethylene biosynthesis
and ethylene
perception genes
during the transition to
climacteric in tomato
Kevany et al 2007
Bleecker and Kende 2000
Ethylene Perception and Signal Transduction
Fruit pulp or the mesocarp
parenchymatus cells
complex network of polysaccharides and proteins
The primary cell wall contains
35 pectin
25 cellulose
20 hemicellulose
10 structural hydroxyproline-rich protein
Structural components of fruits
Plant polysaccharides play a major role in
storage mobilization of energy and in
maintaining cell and tissue integrity due to
their structural and water binding capacity
Enzymes Related to Pectin Dissolution
Critical Reviews in Food Science and Nutrition 471ndash19 (2007)
Globally cultivated fleshy fruit
Worldrsquos largest vegetable crop after potato
Indian production scenario-
350000 hectares 5300000 tonsyear
Short generation time 3-4 months
Simple genetics
Numerous characterized mutants
Cross fertile wild germplasm to promote genetic studies
Routine transformation technology
Postharvest losses-5 to 25 in developed countries
-20 to 50 in developing countries
Tomato model systems for fruit development
and ripening
Natural Mutants Affected in the Ripening Phenotype
Disadvantages of existing methods of storage
Labor intensive
costly
Occupies a large floor space
Poor heat transfer may occur resulting in poor product quality
Excessive dehydration in unpacked products
Chemical changes during freezing
-enzyme-activated browning
-development of rancid oxidative flavors
Textural changes during freezing
-mushy and watery
The use of 1-methylcyclopropene (1-MCP) on fruits and
vegetables
Inhibitor of ethylene perception
Easily released as a gas when the powder is dissolved in water
Approved by the Environmental Protection Agency (EPA) in 1999
Marketed as EthylBlocreg by Floralife Inc (Walterboro SC)AgroFreshInc a
subsidiary of Rohm and Haas (Springhouse PA)
CB WatkinsBiotechnology Advances 24 (2006) 389ndash409
Transgenic approach
Delayed fruit
ripening
BLOCKING THE PERCEPTION OF ETHYLENE
BLOCKING THE EXPRESSION
OF GENES INDUCED BY ETHYLENE
BLOCKING ETHYLENE SYNTHESIS
Regulation of Ethylene Production
a Suppression of ACC synthase gene expression
ACC (1-aminocyclopropane-1-carboxylic acid) (ACS2)
conversion of S-adenosylmethionine (SAM) to ACC
the second to the last step in ethylene biosynthesis
an antisense (ldquomirror-imagerdquo) or truncated copy of the synthase gene
Oeller et al 1991
Yao et al1999
Nath et al 2006
Antisense Technology
httpagbiosafetyunleduflashantisenseswf Journal of Plant Physiology170987ndash 9952013
Null Mutation of the MdACS3 Gene
Apple cultivars homozygous or heterozygous for null allelotype
showed no or very low expression of ripening-related genes and
maintained fruit firmness
Aide Wang 2009
RNAi-mediated silencing
Chimeric RNAi-ACS construct designed to target ACS
homologs
Delayed ripening and extended shelf life for sim45 days
Aarti Gupta Ram Krishna Pal Delayed ripening and improved fruit processing quality in tomato by RNAi-mediated silencing of three
homologs of 1-aminopropane-1-carboxylate synthase gene Journal of Plant Physiology 170 (2013) 987ndash 995
Regulation of Ethylene Production
b Suppression of ACC oxidase gene expression
It catalyzes the oxidation of ACC to ethylene
The last step in the ethylene biosynthetic pathway
Down regulation through anti-sense technology
Hamilton et al 1990
Ye et al 1996
Xiong et al 2003
Ripening in papaya fruit is altered by ACC
oxidase cosuppression
Fig1Map of the construct pKYCPACOO-1 containing the ACC oxidase
fragment cloned in PKYLX80 in the sense orientation The ACC oxidase
fragment is flanked by the CaMV 35S promoter and the RUBISCO
terminator
Fig2 Ethylene production in papaya transgenic fruits
Rodolfo Loacutepez-Goacutemezet alTransgenic Res 1889ndash97 2009
c Insertion of the ACC deaminase gene
Regulation of Ethylene Production
The gene is obtained from Pseudomonas chlororaphis
(a common nonpathogenic soil bacterium)
It converts ACC to a different compound
Reduce the amount of ACC available for ethylene production
90-97 reduced ethylene production
Klee et al1991
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
The molecular mechanisms controlling the ripening of fruit
Open Archive TOULOUSE Archive Ouverte
Increased respiration
Chlorophyll degradation
Biosynthesis of carotenoids anthocyanins essential
oils Flavor and aroma Components
Increased activity of cell wall-degrading enzymes
Transient increase in ethylene production
What are the changes
Major Developmental Changes during Tomato Fruit
Development and Ripening
The Plant Cell Vol 16 S170ndashS180 2004
Based on their respiratory pattern and ethylene biosynthesis
during ripening
VPrasanna et al
Classification of fruits
Pathway for ethylene biosynthesis
Rate limiting step
Critical Reviews in Food Science and Nutrition 471ndash19 2007
The expression of
ethylene biosynthesis
and ethylene
perception genes
during the transition to
climacteric in tomato
Kevany et al 2007
Bleecker and Kende 2000
Ethylene Perception and Signal Transduction
Fruit pulp or the mesocarp
parenchymatus cells
complex network of polysaccharides and proteins
The primary cell wall contains
35 pectin
25 cellulose
20 hemicellulose
10 structural hydroxyproline-rich protein
Structural components of fruits
Plant polysaccharides play a major role in
storage mobilization of energy and in
maintaining cell and tissue integrity due to
their structural and water binding capacity
Enzymes Related to Pectin Dissolution
Critical Reviews in Food Science and Nutrition 471ndash19 (2007)
Globally cultivated fleshy fruit
Worldrsquos largest vegetable crop after potato
Indian production scenario-
350000 hectares 5300000 tonsyear
Short generation time 3-4 months
Simple genetics
Numerous characterized mutants
Cross fertile wild germplasm to promote genetic studies
Routine transformation technology
Postharvest losses-5 to 25 in developed countries
-20 to 50 in developing countries
Tomato model systems for fruit development
and ripening
Natural Mutants Affected in the Ripening Phenotype
Disadvantages of existing methods of storage
Labor intensive
costly
Occupies a large floor space
Poor heat transfer may occur resulting in poor product quality
Excessive dehydration in unpacked products
Chemical changes during freezing
-enzyme-activated browning
-development of rancid oxidative flavors
Textural changes during freezing
-mushy and watery
The use of 1-methylcyclopropene (1-MCP) on fruits and
vegetables
Inhibitor of ethylene perception
Easily released as a gas when the powder is dissolved in water
Approved by the Environmental Protection Agency (EPA) in 1999
Marketed as EthylBlocreg by Floralife Inc (Walterboro SC)AgroFreshInc a
subsidiary of Rohm and Haas (Springhouse PA)
CB WatkinsBiotechnology Advances 24 (2006) 389ndash409
Transgenic approach
Delayed fruit
ripening
BLOCKING THE PERCEPTION OF ETHYLENE
BLOCKING THE EXPRESSION
OF GENES INDUCED BY ETHYLENE
BLOCKING ETHYLENE SYNTHESIS
Regulation of Ethylene Production
a Suppression of ACC synthase gene expression
ACC (1-aminocyclopropane-1-carboxylic acid) (ACS2)
conversion of S-adenosylmethionine (SAM) to ACC
the second to the last step in ethylene biosynthesis
an antisense (ldquomirror-imagerdquo) or truncated copy of the synthase gene
Oeller et al 1991
Yao et al1999
Nath et al 2006
Antisense Technology
httpagbiosafetyunleduflashantisenseswf Journal of Plant Physiology170987ndash 9952013
Null Mutation of the MdACS3 Gene
Apple cultivars homozygous or heterozygous for null allelotype
showed no or very low expression of ripening-related genes and
maintained fruit firmness
Aide Wang 2009
RNAi-mediated silencing
Chimeric RNAi-ACS construct designed to target ACS
homologs
Delayed ripening and extended shelf life for sim45 days
Aarti Gupta Ram Krishna Pal Delayed ripening and improved fruit processing quality in tomato by RNAi-mediated silencing of three
homologs of 1-aminopropane-1-carboxylate synthase gene Journal of Plant Physiology 170 (2013) 987ndash 995
Regulation of Ethylene Production
b Suppression of ACC oxidase gene expression
It catalyzes the oxidation of ACC to ethylene
The last step in the ethylene biosynthetic pathway
Down regulation through anti-sense technology
Hamilton et al 1990
Ye et al 1996
Xiong et al 2003
Ripening in papaya fruit is altered by ACC
oxidase cosuppression
Fig1Map of the construct pKYCPACOO-1 containing the ACC oxidase
fragment cloned in PKYLX80 in the sense orientation The ACC oxidase
fragment is flanked by the CaMV 35S promoter and the RUBISCO
terminator
Fig2 Ethylene production in papaya transgenic fruits
Rodolfo Loacutepez-Goacutemezet alTransgenic Res 1889ndash97 2009
c Insertion of the ACC deaminase gene
Regulation of Ethylene Production
The gene is obtained from Pseudomonas chlororaphis
(a common nonpathogenic soil bacterium)
It converts ACC to a different compound
Reduce the amount of ACC available for ethylene production
90-97 reduced ethylene production
Klee et al1991
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Increased respiration
Chlorophyll degradation
Biosynthesis of carotenoids anthocyanins essential
oils Flavor and aroma Components
Increased activity of cell wall-degrading enzymes
Transient increase in ethylene production
What are the changes
Major Developmental Changes during Tomato Fruit
Development and Ripening
The Plant Cell Vol 16 S170ndashS180 2004
Based on their respiratory pattern and ethylene biosynthesis
during ripening
VPrasanna et al
Classification of fruits
Pathway for ethylene biosynthesis
Rate limiting step
Critical Reviews in Food Science and Nutrition 471ndash19 2007
The expression of
ethylene biosynthesis
and ethylene
perception genes
during the transition to
climacteric in tomato
Kevany et al 2007
Bleecker and Kende 2000
Ethylene Perception and Signal Transduction
Fruit pulp or the mesocarp
parenchymatus cells
complex network of polysaccharides and proteins
The primary cell wall contains
35 pectin
25 cellulose
20 hemicellulose
10 structural hydroxyproline-rich protein
Structural components of fruits
Plant polysaccharides play a major role in
storage mobilization of energy and in
maintaining cell and tissue integrity due to
their structural and water binding capacity
Enzymes Related to Pectin Dissolution
Critical Reviews in Food Science and Nutrition 471ndash19 (2007)
Globally cultivated fleshy fruit
Worldrsquos largest vegetable crop after potato
Indian production scenario-
350000 hectares 5300000 tonsyear
Short generation time 3-4 months
Simple genetics
Numerous characterized mutants
Cross fertile wild germplasm to promote genetic studies
Routine transformation technology
Postharvest losses-5 to 25 in developed countries
-20 to 50 in developing countries
Tomato model systems for fruit development
and ripening
Natural Mutants Affected in the Ripening Phenotype
Disadvantages of existing methods of storage
Labor intensive
costly
Occupies a large floor space
Poor heat transfer may occur resulting in poor product quality
Excessive dehydration in unpacked products
Chemical changes during freezing
-enzyme-activated browning
-development of rancid oxidative flavors
Textural changes during freezing
-mushy and watery
The use of 1-methylcyclopropene (1-MCP) on fruits and
vegetables
Inhibitor of ethylene perception
Easily released as a gas when the powder is dissolved in water
Approved by the Environmental Protection Agency (EPA) in 1999
Marketed as EthylBlocreg by Floralife Inc (Walterboro SC)AgroFreshInc a
subsidiary of Rohm and Haas (Springhouse PA)
CB WatkinsBiotechnology Advances 24 (2006) 389ndash409
Transgenic approach
Delayed fruit
ripening
BLOCKING THE PERCEPTION OF ETHYLENE
BLOCKING THE EXPRESSION
OF GENES INDUCED BY ETHYLENE
BLOCKING ETHYLENE SYNTHESIS
Regulation of Ethylene Production
a Suppression of ACC synthase gene expression
ACC (1-aminocyclopropane-1-carboxylic acid) (ACS2)
conversion of S-adenosylmethionine (SAM) to ACC
the second to the last step in ethylene biosynthesis
an antisense (ldquomirror-imagerdquo) or truncated copy of the synthase gene
Oeller et al 1991
Yao et al1999
Nath et al 2006
Antisense Technology
httpagbiosafetyunleduflashantisenseswf Journal of Plant Physiology170987ndash 9952013
Null Mutation of the MdACS3 Gene
Apple cultivars homozygous or heterozygous for null allelotype
showed no or very low expression of ripening-related genes and
maintained fruit firmness
Aide Wang 2009
RNAi-mediated silencing
Chimeric RNAi-ACS construct designed to target ACS
homologs
Delayed ripening and extended shelf life for sim45 days
Aarti Gupta Ram Krishna Pal Delayed ripening and improved fruit processing quality in tomato by RNAi-mediated silencing of three
homologs of 1-aminopropane-1-carboxylate synthase gene Journal of Plant Physiology 170 (2013) 987ndash 995
Regulation of Ethylene Production
b Suppression of ACC oxidase gene expression
It catalyzes the oxidation of ACC to ethylene
The last step in the ethylene biosynthetic pathway
Down regulation through anti-sense technology
Hamilton et al 1990
Ye et al 1996
Xiong et al 2003
Ripening in papaya fruit is altered by ACC
oxidase cosuppression
Fig1Map of the construct pKYCPACOO-1 containing the ACC oxidase
fragment cloned in PKYLX80 in the sense orientation The ACC oxidase
fragment is flanked by the CaMV 35S promoter and the RUBISCO
terminator
Fig2 Ethylene production in papaya transgenic fruits
Rodolfo Loacutepez-Goacutemezet alTransgenic Res 1889ndash97 2009
c Insertion of the ACC deaminase gene
Regulation of Ethylene Production
The gene is obtained from Pseudomonas chlororaphis
(a common nonpathogenic soil bacterium)
It converts ACC to a different compound
Reduce the amount of ACC available for ethylene production
90-97 reduced ethylene production
Klee et al1991
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Major Developmental Changes during Tomato Fruit
Development and Ripening
The Plant Cell Vol 16 S170ndashS180 2004
Based on their respiratory pattern and ethylene biosynthesis
during ripening
VPrasanna et al
Classification of fruits
Pathway for ethylene biosynthesis
Rate limiting step
Critical Reviews in Food Science and Nutrition 471ndash19 2007
The expression of
ethylene biosynthesis
and ethylene
perception genes
during the transition to
climacteric in tomato
Kevany et al 2007
Bleecker and Kende 2000
Ethylene Perception and Signal Transduction
Fruit pulp or the mesocarp
parenchymatus cells
complex network of polysaccharides and proteins
The primary cell wall contains
35 pectin
25 cellulose
20 hemicellulose
10 structural hydroxyproline-rich protein
Structural components of fruits
Plant polysaccharides play a major role in
storage mobilization of energy and in
maintaining cell and tissue integrity due to
their structural and water binding capacity
Enzymes Related to Pectin Dissolution
Critical Reviews in Food Science and Nutrition 471ndash19 (2007)
Globally cultivated fleshy fruit
Worldrsquos largest vegetable crop after potato
Indian production scenario-
350000 hectares 5300000 tonsyear
Short generation time 3-4 months
Simple genetics
Numerous characterized mutants
Cross fertile wild germplasm to promote genetic studies
Routine transformation technology
Postharvest losses-5 to 25 in developed countries
-20 to 50 in developing countries
Tomato model systems for fruit development
and ripening
Natural Mutants Affected in the Ripening Phenotype
Disadvantages of existing methods of storage
Labor intensive
costly
Occupies a large floor space
Poor heat transfer may occur resulting in poor product quality
Excessive dehydration in unpacked products
Chemical changes during freezing
-enzyme-activated browning
-development of rancid oxidative flavors
Textural changes during freezing
-mushy and watery
The use of 1-methylcyclopropene (1-MCP) on fruits and
vegetables
Inhibitor of ethylene perception
Easily released as a gas when the powder is dissolved in water
Approved by the Environmental Protection Agency (EPA) in 1999
Marketed as EthylBlocreg by Floralife Inc (Walterboro SC)AgroFreshInc a
subsidiary of Rohm and Haas (Springhouse PA)
CB WatkinsBiotechnology Advances 24 (2006) 389ndash409
Transgenic approach
Delayed fruit
ripening
BLOCKING THE PERCEPTION OF ETHYLENE
BLOCKING THE EXPRESSION
OF GENES INDUCED BY ETHYLENE
BLOCKING ETHYLENE SYNTHESIS
Regulation of Ethylene Production
a Suppression of ACC synthase gene expression
ACC (1-aminocyclopropane-1-carboxylic acid) (ACS2)
conversion of S-adenosylmethionine (SAM) to ACC
the second to the last step in ethylene biosynthesis
an antisense (ldquomirror-imagerdquo) or truncated copy of the synthase gene
Oeller et al 1991
Yao et al1999
Nath et al 2006
Antisense Technology
httpagbiosafetyunleduflashantisenseswf Journal of Plant Physiology170987ndash 9952013
Null Mutation of the MdACS3 Gene
Apple cultivars homozygous or heterozygous for null allelotype
showed no or very low expression of ripening-related genes and
maintained fruit firmness
Aide Wang 2009
RNAi-mediated silencing
Chimeric RNAi-ACS construct designed to target ACS
homologs
Delayed ripening and extended shelf life for sim45 days
Aarti Gupta Ram Krishna Pal Delayed ripening and improved fruit processing quality in tomato by RNAi-mediated silencing of three
homologs of 1-aminopropane-1-carboxylate synthase gene Journal of Plant Physiology 170 (2013) 987ndash 995
Regulation of Ethylene Production
b Suppression of ACC oxidase gene expression
It catalyzes the oxidation of ACC to ethylene
The last step in the ethylene biosynthetic pathway
Down regulation through anti-sense technology
Hamilton et al 1990
Ye et al 1996
Xiong et al 2003
Ripening in papaya fruit is altered by ACC
oxidase cosuppression
Fig1Map of the construct pKYCPACOO-1 containing the ACC oxidase
fragment cloned in PKYLX80 in the sense orientation The ACC oxidase
fragment is flanked by the CaMV 35S promoter and the RUBISCO
terminator
Fig2 Ethylene production in papaya transgenic fruits
Rodolfo Loacutepez-Goacutemezet alTransgenic Res 1889ndash97 2009
c Insertion of the ACC deaminase gene
Regulation of Ethylene Production
The gene is obtained from Pseudomonas chlororaphis
(a common nonpathogenic soil bacterium)
It converts ACC to a different compound
Reduce the amount of ACC available for ethylene production
90-97 reduced ethylene production
Klee et al1991
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Based on their respiratory pattern and ethylene biosynthesis
during ripening
VPrasanna et al
Classification of fruits
Pathway for ethylene biosynthesis
Rate limiting step
Critical Reviews in Food Science and Nutrition 471ndash19 2007
The expression of
ethylene biosynthesis
and ethylene
perception genes
during the transition to
climacteric in tomato
Kevany et al 2007
Bleecker and Kende 2000
Ethylene Perception and Signal Transduction
Fruit pulp or the mesocarp
parenchymatus cells
complex network of polysaccharides and proteins
The primary cell wall contains
35 pectin
25 cellulose
20 hemicellulose
10 structural hydroxyproline-rich protein
Structural components of fruits
Plant polysaccharides play a major role in
storage mobilization of energy and in
maintaining cell and tissue integrity due to
their structural and water binding capacity
Enzymes Related to Pectin Dissolution
Critical Reviews in Food Science and Nutrition 471ndash19 (2007)
Globally cultivated fleshy fruit
Worldrsquos largest vegetable crop after potato
Indian production scenario-
350000 hectares 5300000 tonsyear
Short generation time 3-4 months
Simple genetics
Numerous characterized mutants
Cross fertile wild germplasm to promote genetic studies
Routine transformation technology
Postharvest losses-5 to 25 in developed countries
-20 to 50 in developing countries
Tomato model systems for fruit development
and ripening
Natural Mutants Affected in the Ripening Phenotype
Disadvantages of existing methods of storage
Labor intensive
costly
Occupies a large floor space
Poor heat transfer may occur resulting in poor product quality
Excessive dehydration in unpacked products
Chemical changes during freezing
-enzyme-activated browning
-development of rancid oxidative flavors
Textural changes during freezing
-mushy and watery
The use of 1-methylcyclopropene (1-MCP) on fruits and
vegetables
Inhibitor of ethylene perception
Easily released as a gas when the powder is dissolved in water
Approved by the Environmental Protection Agency (EPA) in 1999
Marketed as EthylBlocreg by Floralife Inc (Walterboro SC)AgroFreshInc a
subsidiary of Rohm and Haas (Springhouse PA)
CB WatkinsBiotechnology Advances 24 (2006) 389ndash409
Transgenic approach
Delayed fruit
ripening
BLOCKING THE PERCEPTION OF ETHYLENE
BLOCKING THE EXPRESSION
OF GENES INDUCED BY ETHYLENE
BLOCKING ETHYLENE SYNTHESIS
Regulation of Ethylene Production
a Suppression of ACC synthase gene expression
ACC (1-aminocyclopropane-1-carboxylic acid) (ACS2)
conversion of S-adenosylmethionine (SAM) to ACC
the second to the last step in ethylene biosynthesis
an antisense (ldquomirror-imagerdquo) or truncated copy of the synthase gene
Oeller et al 1991
Yao et al1999
Nath et al 2006
Antisense Technology
httpagbiosafetyunleduflashantisenseswf Journal of Plant Physiology170987ndash 9952013
Null Mutation of the MdACS3 Gene
Apple cultivars homozygous or heterozygous for null allelotype
showed no or very low expression of ripening-related genes and
maintained fruit firmness
Aide Wang 2009
RNAi-mediated silencing
Chimeric RNAi-ACS construct designed to target ACS
homologs
Delayed ripening and extended shelf life for sim45 days
Aarti Gupta Ram Krishna Pal Delayed ripening and improved fruit processing quality in tomato by RNAi-mediated silencing of three
homologs of 1-aminopropane-1-carboxylate synthase gene Journal of Plant Physiology 170 (2013) 987ndash 995
Regulation of Ethylene Production
b Suppression of ACC oxidase gene expression
It catalyzes the oxidation of ACC to ethylene
The last step in the ethylene biosynthetic pathway
Down regulation through anti-sense technology
Hamilton et al 1990
Ye et al 1996
Xiong et al 2003
Ripening in papaya fruit is altered by ACC
oxidase cosuppression
Fig1Map of the construct pKYCPACOO-1 containing the ACC oxidase
fragment cloned in PKYLX80 in the sense orientation The ACC oxidase
fragment is flanked by the CaMV 35S promoter and the RUBISCO
terminator
Fig2 Ethylene production in papaya transgenic fruits
Rodolfo Loacutepez-Goacutemezet alTransgenic Res 1889ndash97 2009
c Insertion of the ACC deaminase gene
Regulation of Ethylene Production
The gene is obtained from Pseudomonas chlororaphis
(a common nonpathogenic soil bacterium)
It converts ACC to a different compound
Reduce the amount of ACC available for ethylene production
90-97 reduced ethylene production
Klee et al1991
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Pathway for ethylene biosynthesis
Rate limiting step
Critical Reviews in Food Science and Nutrition 471ndash19 2007
The expression of
ethylene biosynthesis
and ethylene
perception genes
during the transition to
climacteric in tomato
Kevany et al 2007
Bleecker and Kende 2000
Ethylene Perception and Signal Transduction
Fruit pulp or the mesocarp
parenchymatus cells
complex network of polysaccharides and proteins
The primary cell wall contains
35 pectin
25 cellulose
20 hemicellulose
10 structural hydroxyproline-rich protein
Structural components of fruits
Plant polysaccharides play a major role in
storage mobilization of energy and in
maintaining cell and tissue integrity due to
their structural and water binding capacity
Enzymes Related to Pectin Dissolution
Critical Reviews in Food Science and Nutrition 471ndash19 (2007)
Globally cultivated fleshy fruit
Worldrsquos largest vegetable crop after potato
Indian production scenario-
350000 hectares 5300000 tonsyear
Short generation time 3-4 months
Simple genetics
Numerous characterized mutants
Cross fertile wild germplasm to promote genetic studies
Routine transformation technology
Postharvest losses-5 to 25 in developed countries
-20 to 50 in developing countries
Tomato model systems for fruit development
and ripening
Natural Mutants Affected in the Ripening Phenotype
Disadvantages of existing methods of storage
Labor intensive
costly
Occupies a large floor space
Poor heat transfer may occur resulting in poor product quality
Excessive dehydration in unpacked products
Chemical changes during freezing
-enzyme-activated browning
-development of rancid oxidative flavors
Textural changes during freezing
-mushy and watery
The use of 1-methylcyclopropene (1-MCP) on fruits and
vegetables
Inhibitor of ethylene perception
Easily released as a gas when the powder is dissolved in water
Approved by the Environmental Protection Agency (EPA) in 1999
Marketed as EthylBlocreg by Floralife Inc (Walterboro SC)AgroFreshInc a
subsidiary of Rohm and Haas (Springhouse PA)
CB WatkinsBiotechnology Advances 24 (2006) 389ndash409
Transgenic approach
Delayed fruit
ripening
BLOCKING THE PERCEPTION OF ETHYLENE
BLOCKING THE EXPRESSION
OF GENES INDUCED BY ETHYLENE
BLOCKING ETHYLENE SYNTHESIS
Regulation of Ethylene Production
a Suppression of ACC synthase gene expression
ACC (1-aminocyclopropane-1-carboxylic acid) (ACS2)
conversion of S-adenosylmethionine (SAM) to ACC
the second to the last step in ethylene biosynthesis
an antisense (ldquomirror-imagerdquo) or truncated copy of the synthase gene
Oeller et al 1991
Yao et al1999
Nath et al 2006
Antisense Technology
httpagbiosafetyunleduflashantisenseswf Journal of Plant Physiology170987ndash 9952013
Null Mutation of the MdACS3 Gene
Apple cultivars homozygous or heterozygous for null allelotype
showed no or very low expression of ripening-related genes and
maintained fruit firmness
Aide Wang 2009
RNAi-mediated silencing
Chimeric RNAi-ACS construct designed to target ACS
homologs
Delayed ripening and extended shelf life for sim45 days
Aarti Gupta Ram Krishna Pal Delayed ripening and improved fruit processing quality in tomato by RNAi-mediated silencing of three
homologs of 1-aminopropane-1-carboxylate synthase gene Journal of Plant Physiology 170 (2013) 987ndash 995
Regulation of Ethylene Production
b Suppression of ACC oxidase gene expression
It catalyzes the oxidation of ACC to ethylene
The last step in the ethylene biosynthetic pathway
Down regulation through anti-sense technology
Hamilton et al 1990
Ye et al 1996
Xiong et al 2003
Ripening in papaya fruit is altered by ACC
oxidase cosuppression
Fig1Map of the construct pKYCPACOO-1 containing the ACC oxidase
fragment cloned in PKYLX80 in the sense orientation The ACC oxidase
fragment is flanked by the CaMV 35S promoter and the RUBISCO
terminator
Fig2 Ethylene production in papaya transgenic fruits
Rodolfo Loacutepez-Goacutemezet alTransgenic Res 1889ndash97 2009
c Insertion of the ACC deaminase gene
Regulation of Ethylene Production
The gene is obtained from Pseudomonas chlororaphis
(a common nonpathogenic soil bacterium)
It converts ACC to a different compound
Reduce the amount of ACC available for ethylene production
90-97 reduced ethylene production
Klee et al1991
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
The expression of
ethylene biosynthesis
and ethylene
perception genes
during the transition to
climacteric in tomato
Kevany et al 2007
Bleecker and Kende 2000
Ethylene Perception and Signal Transduction
Fruit pulp or the mesocarp
parenchymatus cells
complex network of polysaccharides and proteins
The primary cell wall contains
35 pectin
25 cellulose
20 hemicellulose
10 structural hydroxyproline-rich protein
Structural components of fruits
Plant polysaccharides play a major role in
storage mobilization of energy and in
maintaining cell and tissue integrity due to
their structural and water binding capacity
Enzymes Related to Pectin Dissolution
Critical Reviews in Food Science and Nutrition 471ndash19 (2007)
Globally cultivated fleshy fruit
Worldrsquos largest vegetable crop after potato
Indian production scenario-
350000 hectares 5300000 tonsyear
Short generation time 3-4 months
Simple genetics
Numerous characterized mutants
Cross fertile wild germplasm to promote genetic studies
Routine transformation technology
Postharvest losses-5 to 25 in developed countries
-20 to 50 in developing countries
Tomato model systems for fruit development
and ripening
Natural Mutants Affected in the Ripening Phenotype
Disadvantages of existing methods of storage
Labor intensive
costly
Occupies a large floor space
Poor heat transfer may occur resulting in poor product quality
Excessive dehydration in unpacked products
Chemical changes during freezing
-enzyme-activated browning
-development of rancid oxidative flavors
Textural changes during freezing
-mushy and watery
The use of 1-methylcyclopropene (1-MCP) on fruits and
vegetables
Inhibitor of ethylene perception
Easily released as a gas when the powder is dissolved in water
Approved by the Environmental Protection Agency (EPA) in 1999
Marketed as EthylBlocreg by Floralife Inc (Walterboro SC)AgroFreshInc a
subsidiary of Rohm and Haas (Springhouse PA)
CB WatkinsBiotechnology Advances 24 (2006) 389ndash409
Transgenic approach
Delayed fruit
ripening
BLOCKING THE PERCEPTION OF ETHYLENE
BLOCKING THE EXPRESSION
OF GENES INDUCED BY ETHYLENE
BLOCKING ETHYLENE SYNTHESIS
Regulation of Ethylene Production
a Suppression of ACC synthase gene expression
ACC (1-aminocyclopropane-1-carboxylic acid) (ACS2)
conversion of S-adenosylmethionine (SAM) to ACC
the second to the last step in ethylene biosynthesis
an antisense (ldquomirror-imagerdquo) or truncated copy of the synthase gene
Oeller et al 1991
Yao et al1999
Nath et al 2006
Antisense Technology
httpagbiosafetyunleduflashantisenseswf Journal of Plant Physiology170987ndash 9952013
Null Mutation of the MdACS3 Gene
Apple cultivars homozygous or heterozygous for null allelotype
showed no or very low expression of ripening-related genes and
maintained fruit firmness
Aide Wang 2009
RNAi-mediated silencing
Chimeric RNAi-ACS construct designed to target ACS
homologs
Delayed ripening and extended shelf life for sim45 days
Aarti Gupta Ram Krishna Pal Delayed ripening and improved fruit processing quality in tomato by RNAi-mediated silencing of three
homologs of 1-aminopropane-1-carboxylate synthase gene Journal of Plant Physiology 170 (2013) 987ndash 995
Regulation of Ethylene Production
b Suppression of ACC oxidase gene expression
It catalyzes the oxidation of ACC to ethylene
The last step in the ethylene biosynthetic pathway
Down regulation through anti-sense technology
Hamilton et al 1990
Ye et al 1996
Xiong et al 2003
Ripening in papaya fruit is altered by ACC
oxidase cosuppression
Fig1Map of the construct pKYCPACOO-1 containing the ACC oxidase
fragment cloned in PKYLX80 in the sense orientation The ACC oxidase
fragment is flanked by the CaMV 35S promoter and the RUBISCO
terminator
Fig2 Ethylene production in papaya transgenic fruits
Rodolfo Loacutepez-Goacutemezet alTransgenic Res 1889ndash97 2009
c Insertion of the ACC deaminase gene
Regulation of Ethylene Production
The gene is obtained from Pseudomonas chlororaphis
(a common nonpathogenic soil bacterium)
It converts ACC to a different compound
Reduce the amount of ACC available for ethylene production
90-97 reduced ethylene production
Klee et al1991
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Bleecker and Kende 2000
Ethylene Perception and Signal Transduction
Fruit pulp or the mesocarp
parenchymatus cells
complex network of polysaccharides and proteins
The primary cell wall contains
35 pectin
25 cellulose
20 hemicellulose
10 structural hydroxyproline-rich protein
Structural components of fruits
Plant polysaccharides play a major role in
storage mobilization of energy and in
maintaining cell and tissue integrity due to
their structural and water binding capacity
Enzymes Related to Pectin Dissolution
Critical Reviews in Food Science and Nutrition 471ndash19 (2007)
Globally cultivated fleshy fruit
Worldrsquos largest vegetable crop after potato
Indian production scenario-
350000 hectares 5300000 tonsyear
Short generation time 3-4 months
Simple genetics
Numerous characterized mutants
Cross fertile wild germplasm to promote genetic studies
Routine transformation technology
Postharvest losses-5 to 25 in developed countries
-20 to 50 in developing countries
Tomato model systems for fruit development
and ripening
Natural Mutants Affected in the Ripening Phenotype
Disadvantages of existing methods of storage
Labor intensive
costly
Occupies a large floor space
Poor heat transfer may occur resulting in poor product quality
Excessive dehydration in unpacked products
Chemical changes during freezing
-enzyme-activated browning
-development of rancid oxidative flavors
Textural changes during freezing
-mushy and watery
The use of 1-methylcyclopropene (1-MCP) on fruits and
vegetables
Inhibitor of ethylene perception
Easily released as a gas when the powder is dissolved in water
Approved by the Environmental Protection Agency (EPA) in 1999
Marketed as EthylBlocreg by Floralife Inc (Walterboro SC)AgroFreshInc a
subsidiary of Rohm and Haas (Springhouse PA)
CB WatkinsBiotechnology Advances 24 (2006) 389ndash409
Transgenic approach
Delayed fruit
ripening
BLOCKING THE PERCEPTION OF ETHYLENE
BLOCKING THE EXPRESSION
OF GENES INDUCED BY ETHYLENE
BLOCKING ETHYLENE SYNTHESIS
Regulation of Ethylene Production
a Suppression of ACC synthase gene expression
ACC (1-aminocyclopropane-1-carboxylic acid) (ACS2)
conversion of S-adenosylmethionine (SAM) to ACC
the second to the last step in ethylene biosynthesis
an antisense (ldquomirror-imagerdquo) or truncated copy of the synthase gene
Oeller et al 1991
Yao et al1999
Nath et al 2006
Antisense Technology
httpagbiosafetyunleduflashantisenseswf Journal of Plant Physiology170987ndash 9952013
Null Mutation of the MdACS3 Gene
Apple cultivars homozygous or heterozygous for null allelotype
showed no or very low expression of ripening-related genes and
maintained fruit firmness
Aide Wang 2009
RNAi-mediated silencing
Chimeric RNAi-ACS construct designed to target ACS
homologs
Delayed ripening and extended shelf life for sim45 days
Aarti Gupta Ram Krishna Pal Delayed ripening and improved fruit processing quality in tomato by RNAi-mediated silencing of three
homologs of 1-aminopropane-1-carboxylate synthase gene Journal of Plant Physiology 170 (2013) 987ndash 995
Regulation of Ethylene Production
b Suppression of ACC oxidase gene expression
It catalyzes the oxidation of ACC to ethylene
The last step in the ethylene biosynthetic pathway
Down regulation through anti-sense technology
Hamilton et al 1990
Ye et al 1996
Xiong et al 2003
Ripening in papaya fruit is altered by ACC
oxidase cosuppression
Fig1Map of the construct pKYCPACOO-1 containing the ACC oxidase
fragment cloned in PKYLX80 in the sense orientation The ACC oxidase
fragment is flanked by the CaMV 35S promoter and the RUBISCO
terminator
Fig2 Ethylene production in papaya transgenic fruits
Rodolfo Loacutepez-Goacutemezet alTransgenic Res 1889ndash97 2009
c Insertion of the ACC deaminase gene
Regulation of Ethylene Production
The gene is obtained from Pseudomonas chlororaphis
(a common nonpathogenic soil bacterium)
It converts ACC to a different compound
Reduce the amount of ACC available for ethylene production
90-97 reduced ethylene production
Klee et al1991
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Fruit pulp or the mesocarp
parenchymatus cells
complex network of polysaccharides and proteins
The primary cell wall contains
35 pectin
25 cellulose
20 hemicellulose
10 structural hydroxyproline-rich protein
Structural components of fruits
Plant polysaccharides play a major role in
storage mobilization of energy and in
maintaining cell and tissue integrity due to
their structural and water binding capacity
Enzymes Related to Pectin Dissolution
Critical Reviews in Food Science and Nutrition 471ndash19 (2007)
Globally cultivated fleshy fruit
Worldrsquos largest vegetable crop after potato
Indian production scenario-
350000 hectares 5300000 tonsyear
Short generation time 3-4 months
Simple genetics
Numerous characterized mutants
Cross fertile wild germplasm to promote genetic studies
Routine transformation technology
Postharvest losses-5 to 25 in developed countries
-20 to 50 in developing countries
Tomato model systems for fruit development
and ripening
Natural Mutants Affected in the Ripening Phenotype
Disadvantages of existing methods of storage
Labor intensive
costly
Occupies a large floor space
Poor heat transfer may occur resulting in poor product quality
Excessive dehydration in unpacked products
Chemical changes during freezing
-enzyme-activated browning
-development of rancid oxidative flavors
Textural changes during freezing
-mushy and watery
The use of 1-methylcyclopropene (1-MCP) on fruits and
vegetables
Inhibitor of ethylene perception
Easily released as a gas when the powder is dissolved in water
Approved by the Environmental Protection Agency (EPA) in 1999
Marketed as EthylBlocreg by Floralife Inc (Walterboro SC)AgroFreshInc a
subsidiary of Rohm and Haas (Springhouse PA)
CB WatkinsBiotechnology Advances 24 (2006) 389ndash409
Transgenic approach
Delayed fruit
ripening
BLOCKING THE PERCEPTION OF ETHYLENE
BLOCKING THE EXPRESSION
OF GENES INDUCED BY ETHYLENE
BLOCKING ETHYLENE SYNTHESIS
Regulation of Ethylene Production
a Suppression of ACC synthase gene expression
ACC (1-aminocyclopropane-1-carboxylic acid) (ACS2)
conversion of S-adenosylmethionine (SAM) to ACC
the second to the last step in ethylene biosynthesis
an antisense (ldquomirror-imagerdquo) or truncated copy of the synthase gene
Oeller et al 1991
Yao et al1999
Nath et al 2006
Antisense Technology
httpagbiosafetyunleduflashantisenseswf Journal of Plant Physiology170987ndash 9952013
Null Mutation of the MdACS3 Gene
Apple cultivars homozygous or heterozygous for null allelotype
showed no or very low expression of ripening-related genes and
maintained fruit firmness
Aide Wang 2009
RNAi-mediated silencing
Chimeric RNAi-ACS construct designed to target ACS
homologs
Delayed ripening and extended shelf life for sim45 days
Aarti Gupta Ram Krishna Pal Delayed ripening and improved fruit processing quality in tomato by RNAi-mediated silencing of three
homologs of 1-aminopropane-1-carboxylate synthase gene Journal of Plant Physiology 170 (2013) 987ndash 995
Regulation of Ethylene Production
b Suppression of ACC oxidase gene expression
It catalyzes the oxidation of ACC to ethylene
The last step in the ethylene biosynthetic pathway
Down regulation through anti-sense technology
Hamilton et al 1990
Ye et al 1996
Xiong et al 2003
Ripening in papaya fruit is altered by ACC
oxidase cosuppression
Fig1Map of the construct pKYCPACOO-1 containing the ACC oxidase
fragment cloned in PKYLX80 in the sense orientation The ACC oxidase
fragment is flanked by the CaMV 35S promoter and the RUBISCO
terminator
Fig2 Ethylene production in papaya transgenic fruits
Rodolfo Loacutepez-Goacutemezet alTransgenic Res 1889ndash97 2009
c Insertion of the ACC deaminase gene
Regulation of Ethylene Production
The gene is obtained from Pseudomonas chlororaphis
(a common nonpathogenic soil bacterium)
It converts ACC to a different compound
Reduce the amount of ACC available for ethylene production
90-97 reduced ethylene production
Klee et al1991
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Enzymes Related to Pectin Dissolution
Critical Reviews in Food Science and Nutrition 471ndash19 (2007)
Globally cultivated fleshy fruit
Worldrsquos largest vegetable crop after potato
Indian production scenario-
350000 hectares 5300000 tonsyear
Short generation time 3-4 months
Simple genetics
Numerous characterized mutants
Cross fertile wild germplasm to promote genetic studies
Routine transformation technology
Postharvest losses-5 to 25 in developed countries
-20 to 50 in developing countries
Tomato model systems for fruit development
and ripening
Natural Mutants Affected in the Ripening Phenotype
Disadvantages of existing methods of storage
Labor intensive
costly
Occupies a large floor space
Poor heat transfer may occur resulting in poor product quality
Excessive dehydration in unpacked products
Chemical changes during freezing
-enzyme-activated browning
-development of rancid oxidative flavors
Textural changes during freezing
-mushy and watery
The use of 1-methylcyclopropene (1-MCP) on fruits and
vegetables
Inhibitor of ethylene perception
Easily released as a gas when the powder is dissolved in water
Approved by the Environmental Protection Agency (EPA) in 1999
Marketed as EthylBlocreg by Floralife Inc (Walterboro SC)AgroFreshInc a
subsidiary of Rohm and Haas (Springhouse PA)
CB WatkinsBiotechnology Advances 24 (2006) 389ndash409
Transgenic approach
Delayed fruit
ripening
BLOCKING THE PERCEPTION OF ETHYLENE
BLOCKING THE EXPRESSION
OF GENES INDUCED BY ETHYLENE
BLOCKING ETHYLENE SYNTHESIS
Regulation of Ethylene Production
a Suppression of ACC synthase gene expression
ACC (1-aminocyclopropane-1-carboxylic acid) (ACS2)
conversion of S-adenosylmethionine (SAM) to ACC
the second to the last step in ethylene biosynthesis
an antisense (ldquomirror-imagerdquo) or truncated copy of the synthase gene
Oeller et al 1991
Yao et al1999
Nath et al 2006
Antisense Technology
httpagbiosafetyunleduflashantisenseswf Journal of Plant Physiology170987ndash 9952013
Null Mutation of the MdACS3 Gene
Apple cultivars homozygous or heterozygous for null allelotype
showed no or very low expression of ripening-related genes and
maintained fruit firmness
Aide Wang 2009
RNAi-mediated silencing
Chimeric RNAi-ACS construct designed to target ACS
homologs
Delayed ripening and extended shelf life for sim45 days
Aarti Gupta Ram Krishna Pal Delayed ripening and improved fruit processing quality in tomato by RNAi-mediated silencing of three
homologs of 1-aminopropane-1-carboxylate synthase gene Journal of Plant Physiology 170 (2013) 987ndash 995
Regulation of Ethylene Production
b Suppression of ACC oxidase gene expression
It catalyzes the oxidation of ACC to ethylene
The last step in the ethylene biosynthetic pathway
Down regulation through anti-sense technology
Hamilton et al 1990
Ye et al 1996
Xiong et al 2003
Ripening in papaya fruit is altered by ACC
oxidase cosuppression
Fig1Map of the construct pKYCPACOO-1 containing the ACC oxidase
fragment cloned in PKYLX80 in the sense orientation The ACC oxidase
fragment is flanked by the CaMV 35S promoter and the RUBISCO
terminator
Fig2 Ethylene production in papaya transgenic fruits
Rodolfo Loacutepez-Goacutemezet alTransgenic Res 1889ndash97 2009
c Insertion of the ACC deaminase gene
Regulation of Ethylene Production
The gene is obtained from Pseudomonas chlororaphis
(a common nonpathogenic soil bacterium)
It converts ACC to a different compound
Reduce the amount of ACC available for ethylene production
90-97 reduced ethylene production
Klee et al1991
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Globally cultivated fleshy fruit
Worldrsquos largest vegetable crop after potato
Indian production scenario-
350000 hectares 5300000 tonsyear
Short generation time 3-4 months
Simple genetics
Numerous characterized mutants
Cross fertile wild germplasm to promote genetic studies
Routine transformation technology
Postharvest losses-5 to 25 in developed countries
-20 to 50 in developing countries
Tomato model systems for fruit development
and ripening
Natural Mutants Affected in the Ripening Phenotype
Disadvantages of existing methods of storage
Labor intensive
costly
Occupies a large floor space
Poor heat transfer may occur resulting in poor product quality
Excessive dehydration in unpacked products
Chemical changes during freezing
-enzyme-activated browning
-development of rancid oxidative flavors
Textural changes during freezing
-mushy and watery
The use of 1-methylcyclopropene (1-MCP) on fruits and
vegetables
Inhibitor of ethylene perception
Easily released as a gas when the powder is dissolved in water
Approved by the Environmental Protection Agency (EPA) in 1999
Marketed as EthylBlocreg by Floralife Inc (Walterboro SC)AgroFreshInc a
subsidiary of Rohm and Haas (Springhouse PA)
CB WatkinsBiotechnology Advances 24 (2006) 389ndash409
Transgenic approach
Delayed fruit
ripening
BLOCKING THE PERCEPTION OF ETHYLENE
BLOCKING THE EXPRESSION
OF GENES INDUCED BY ETHYLENE
BLOCKING ETHYLENE SYNTHESIS
Regulation of Ethylene Production
a Suppression of ACC synthase gene expression
ACC (1-aminocyclopropane-1-carboxylic acid) (ACS2)
conversion of S-adenosylmethionine (SAM) to ACC
the second to the last step in ethylene biosynthesis
an antisense (ldquomirror-imagerdquo) or truncated copy of the synthase gene
Oeller et al 1991
Yao et al1999
Nath et al 2006
Antisense Technology
httpagbiosafetyunleduflashantisenseswf Journal of Plant Physiology170987ndash 9952013
Null Mutation of the MdACS3 Gene
Apple cultivars homozygous or heterozygous for null allelotype
showed no or very low expression of ripening-related genes and
maintained fruit firmness
Aide Wang 2009
RNAi-mediated silencing
Chimeric RNAi-ACS construct designed to target ACS
homologs
Delayed ripening and extended shelf life for sim45 days
Aarti Gupta Ram Krishna Pal Delayed ripening and improved fruit processing quality in tomato by RNAi-mediated silencing of three
homologs of 1-aminopropane-1-carboxylate synthase gene Journal of Plant Physiology 170 (2013) 987ndash 995
Regulation of Ethylene Production
b Suppression of ACC oxidase gene expression
It catalyzes the oxidation of ACC to ethylene
The last step in the ethylene biosynthetic pathway
Down regulation through anti-sense technology
Hamilton et al 1990
Ye et al 1996
Xiong et al 2003
Ripening in papaya fruit is altered by ACC
oxidase cosuppression
Fig1Map of the construct pKYCPACOO-1 containing the ACC oxidase
fragment cloned in PKYLX80 in the sense orientation The ACC oxidase
fragment is flanked by the CaMV 35S promoter and the RUBISCO
terminator
Fig2 Ethylene production in papaya transgenic fruits
Rodolfo Loacutepez-Goacutemezet alTransgenic Res 1889ndash97 2009
c Insertion of the ACC deaminase gene
Regulation of Ethylene Production
The gene is obtained from Pseudomonas chlororaphis
(a common nonpathogenic soil bacterium)
It converts ACC to a different compound
Reduce the amount of ACC available for ethylene production
90-97 reduced ethylene production
Klee et al1991
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Natural Mutants Affected in the Ripening Phenotype
Disadvantages of existing methods of storage
Labor intensive
costly
Occupies a large floor space
Poor heat transfer may occur resulting in poor product quality
Excessive dehydration in unpacked products
Chemical changes during freezing
-enzyme-activated browning
-development of rancid oxidative flavors
Textural changes during freezing
-mushy and watery
The use of 1-methylcyclopropene (1-MCP) on fruits and
vegetables
Inhibitor of ethylene perception
Easily released as a gas when the powder is dissolved in water
Approved by the Environmental Protection Agency (EPA) in 1999
Marketed as EthylBlocreg by Floralife Inc (Walterboro SC)AgroFreshInc a
subsidiary of Rohm and Haas (Springhouse PA)
CB WatkinsBiotechnology Advances 24 (2006) 389ndash409
Transgenic approach
Delayed fruit
ripening
BLOCKING THE PERCEPTION OF ETHYLENE
BLOCKING THE EXPRESSION
OF GENES INDUCED BY ETHYLENE
BLOCKING ETHYLENE SYNTHESIS
Regulation of Ethylene Production
a Suppression of ACC synthase gene expression
ACC (1-aminocyclopropane-1-carboxylic acid) (ACS2)
conversion of S-adenosylmethionine (SAM) to ACC
the second to the last step in ethylene biosynthesis
an antisense (ldquomirror-imagerdquo) or truncated copy of the synthase gene
Oeller et al 1991
Yao et al1999
Nath et al 2006
Antisense Technology
httpagbiosafetyunleduflashantisenseswf Journal of Plant Physiology170987ndash 9952013
Null Mutation of the MdACS3 Gene
Apple cultivars homozygous or heterozygous for null allelotype
showed no or very low expression of ripening-related genes and
maintained fruit firmness
Aide Wang 2009
RNAi-mediated silencing
Chimeric RNAi-ACS construct designed to target ACS
homologs
Delayed ripening and extended shelf life for sim45 days
Aarti Gupta Ram Krishna Pal Delayed ripening and improved fruit processing quality in tomato by RNAi-mediated silencing of three
homologs of 1-aminopropane-1-carboxylate synthase gene Journal of Plant Physiology 170 (2013) 987ndash 995
Regulation of Ethylene Production
b Suppression of ACC oxidase gene expression
It catalyzes the oxidation of ACC to ethylene
The last step in the ethylene biosynthetic pathway
Down regulation through anti-sense technology
Hamilton et al 1990
Ye et al 1996
Xiong et al 2003
Ripening in papaya fruit is altered by ACC
oxidase cosuppression
Fig1Map of the construct pKYCPACOO-1 containing the ACC oxidase
fragment cloned in PKYLX80 in the sense orientation The ACC oxidase
fragment is flanked by the CaMV 35S promoter and the RUBISCO
terminator
Fig2 Ethylene production in papaya transgenic fruits
Rodolfo Loacutepez-Goacutemezet alTransgenic Res 1889ndash97 2009
c Insertion of the ACC deaminase gene
Regulation of Ethylene Production
The gene is obtained from Pseudomonas chlororaphis
(a common nonpathogenic soil bacterium)
It converts ACC to a different compound
Reduce the amount of ACC available for ethylene production
90-97 reduced ethylene production
Klee et al1991
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Disadvantages of existing methods of storage
Labor intensive
costly
Occupies a large floor space
Poor heat transfer may occur resulting in poor product quality
Excessive dehydration in unpacked products
Chemical changes during freezing
-enzyme-activated browning
-development of rancid oxidative flavors
Textural changes during freezing
-mushy and watery
The use of 1-methylcyclopropene (1-MCP) on fruits and
vegetables
Inhibitor of ethylene perception
Easily released as a gas when the powder is dissolved in water
Approved by the Environmental Protection Agency (EPA) in 1999
Marketed as EthylBlocreg by Floralife Inc (Walterboro SC)AgroFreshInc a
subsidiary of Rohm and Haas (Springhouse PA)
CB WatkinsBiotechnology Advances 24 (2006) 389ndash409
Transgenic approach
Delayed fruit
ripening
BLOCKING THE PERCEPTION OF ETHYLENE
BLOCKING THE EXPRESSION
OF GENES INDUCED BY ETHYLENE
BLOCKING ETHYLENE SYNTHESIS
Regulation of Ethylene Production
a Suppression of ACC synthase gene expression
ACC (1-aminocyclopropane-1-carboxylic acid) (ACS2)
conversion of S-adenosylmethionine (SAM) to ACC
the second to the last step in ethylene biosynthesis
an antisense (ldquomirror-imagerdquo) or truncated copy of the synthase gene
Oeller et al 1991
Yao et al1999
Nath et al 2006
Antisense Technology
httpagbiosafetyunleduflashantisenseswf Journal of Plant Physiology170987ndash 9952013
Null Mutation of the MdACS3 Gene
Apple cultivars homozygous or heterozygous for null allelotype
showed no or very low expression of ripening-related genes and
maintained fruit firmness
Aide Wang 2009
RNAi-mediated silencing
Chimeric RNAi-ACS construct designed to target ACS
homologs
Delayed ripening and extended shelf life for sim45 days
Aarti Gupta Ram Krishna Pal Delayed ripening and improved fruit processing quality in tomato by RNAi-mediated silencing of three
homologs of 1-aminopropane-1-carboxylate synthase gene Journal of Plant Physiology 170 (2013) 987ndash 995
Regulation of Ethylene Production
b Suppression of ACC oxidase gene expression
It catalyzes the oxidation of ACC to ethylene
The last step in the ethylene biosynthetic pathway
Down regulation through anti-sense technology
Hamilton et al 1990
Ye et al 1996
Xiong et al 2003
Ripening in papaya fruit is altered by ACC
oxidase cosuppression
Fig1Map of the construct pKYCPACOO-1 containing the ACC oxidase
fragment cloned in PKYLX80 in the sense orientation The ACC oxidase
fragment is flanked by the CaMV 35S promoter and the RUBISCO
terminator
Fig2 Ethylene production in papaya transgenic fruits
Rodolfo Loacutepez-Goacutemezet alTransgenic Res 1889ndash97 2009
c Insertion of the ACC deaminase gene
Regulation of Ethylene Production
The gene is obtained from Pseudomonas chlororaphis
(a common nonpathogenic soil bacterium)
It converts ACC to a different compound
Reduce the amount of ACC available for ethylene production
90-97 reduced ethylene production
Klee et al1991
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
The use of 1-methylcyclopropene (1-MCP) on fruits and
vegetables
Inhibitor of ethylene perception
Easily released as a gas when the powder is dissolved in water
Approved by the Environmental Protection Agency (EPA) in 1999
Marketed as EthylBlocreg by Floralife Inc (Walterboro SC)AgroFreshInc a
subsidiary of Rohm and Haas (Springhouse PA)
CB WatkinsBiotechnology Advances 24 (2006) 389ndash409
Transgenic approach
Delayed fruit
ripening
BLOCKING THE PERCEPTION OF ETHYLENE
BLOCKING THE EXPRESSION
OF GENES INDUCED BY ETHYLENE
BLOCKING ETHYLENE SYNTHESIS
Regulation of Ethylene Production
a Suppression of ACC synthase gene expression
ACC (1-aminocyclopropane-1-carboxylic acid) (ACS2)
conversion of S-adenosylmethionine (SAM) to ACC
the second to the last step in ethylene biosynthesis
an antisense (ldquomirror-imagerdquo) or truncated copy of the synthase gene
Oeller et al 1991
Yao et al1999
Nath et al 2006
Antisense Technology
httpagbiosafetyunleduflashantisenseswf Journal of Plant Physiology170987ndash 9952013
Null Mutation of the MdACS3 Gene
Apple cultivars homozygous or heterozygous for null allelotype
showed no or very low expression of ripening-related genes and
maintained fruit firmness
Aide Wang 2009
RNAi-mediated silencing
Chimeric RNAi-ACS construct designed to target ACS
homologs
Delayed ripening and extended shelf life for sim45 days
Aarti Gupta Ram Krishna Pal Delayed ripening and improved fruit processing quality in tomato by RNAi-mediated silencing of three
homologs of 1-aminopropane-1-carboxylate synthase gene Journal of Plant Physiology 170 (2013) 987ndash 995
Regulation of Ethylene Production
b Suppression of ACC oxidase gene expression
It catalyzes the oxidation of ACC to ethylene
The last step in the ethylene biosynthetic pathway
Down regulation through anti-sense technology
Hamilton et al 1990
Ye et al 1996
Xiong et al 2003
Ripening in papaya fruit is altered by ACC
oxidase cosuppression
Fig1Map of the construct pKYCPACOO-1 containing the ACC oxidase
fragment cloned in PKYLX80 in the sense orientation The ACC oxidase
fragment is flanked by the CaMV 35S promoter and the RUBISCO
terminator
Fig2 Ethylene production in papaya transgenic fruits
Rodolfo Loacutepez-Goacutemezet alTransgenic Res 1889ndash97 2009
c Insertion of the ACC deaminase gene
Regulation of Ethylene Production
The gene is obtained from Pseudomonas chlororaphis
(a common nonpathogenic soil bacterium)
It converts ACC to a different compound
Reduce the amount of ACC available for ethylene production
90-97 reduced ethylene production
Klee et al1991
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Transgenic approach
Delayed fruit
ripening
BLOCKING THE PERCEPTION OF ETHYLENE
BLOCKING THE EXPRESSION
OF GENES INDUCED BY ETHYLENE
BLOCKING ETHYLENE SYNTHESIS
Regulation of Ethylene Production
a Suppression of ACC synthase gene expression
ACC (1-aminocyclopropane-1-carboxylic acid) (ACS2)
conversion of S-adenosylmethionine (SAM) to ACC
the second to the last step in ethylene biosynthesis
an antisense (ldquomirror-imagerdquo) or truncated copy of the synthase gene
Oeller et al 1991
Yao et al1999
Nath et al 2006
Antisense Technology
httpagbiosafetyunleduflashantisenseswf Journal of Plant Physiology170987ndash 9952013
Null Mutation of the MdACS3 Gene
Apple cultivars homozygous or heterozygous for null allelotype
showed no or very low expression of ripening-related genes and
maintained fruit firmness
Aide Wang 2009
RNAi-mediated silencing
Chimeric RNAi-ACS construct designed to target ACS
homologs
Delayed ripening and extended shelf life for sim45 days
Aarti Gupta Ram Krishna Pal Delayed ripening and improved fruit processing quality in tomato by RNAi-mediated silencing of three
homologs of 1-aminopropane-1-carboxylate synthase gene Journal of Plant Physiology 170 (2013) 987ndash 995
Regulation of Ethylene Production
b Suppression of ACC oxidase gene expression
It catalyzes the oxidation of ACC to ethylene
The last step in the ethylene biosynthetic pathway
Down regulation through anti-sense technology
Hamilton et al 1990
Ye et al 1996
Xiong et al 2003
Ripening in papaya fruit is altered by ACC
oxidase cosuppression
Fig1Map of the construct pKYCPACOO-1 containing the ACC oxidase
fragment cloned in PKYLX80 in the sense orientation The ACC oxidase
fragment is flanked by the CaMV 35S promoter and the RUBISCO
terminator
Fig2 Ethylene production in papaya transgenic fruits
Rodolfo Loacutepez-Goacutemezet alTransgenic Res 1889ndash97 2009
c Insertion of the ACC deaminase gene
Regulation of Ethylene Production
The gene is obtained from Pseudomonas chlororaphis
(a common nonpathogenic soil bacterium)
It converts ACC to a different compound
Reduce the amount of ACC available for ethylene production
90-97 reduced ethylene production
Klee et al1991
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Regulation of Ethylene Production
a Suppression of ACC synthase gene expression
ACC (1-aminocyclopropane-1-carboxylic acid) (ACS2)
conversion of S-adenosylmethionine (SAM) to ACC
the second to the last step in ethylene biosynthesis
an antisense (ldquomirror-imagerdquo) or truncated copy of the synthase gene
Oeller et al 1991
Yao et al1999
Nath et al 2006
Antisense Technology
httpagbiosafetyunleduflashantisenseswf Journal of Plant Physiology170987ndash 9952013
Null Mutation of the MdACS3 Gene
Apple cultivars homozygous or heterozygous for null allelotype
showed no or very low expression of ripening-related genes and
maintained fruit firmness
Aide Wang 2009
RNAi-mediated silencing
Chimeric RNAi-ACS construct designed to target ACS
homologs
Delayed ripening and extended shelf life for sim45 days
Aarti Gupta Ram Krishna Pal Delayed ripening and improved fruit processing quality in tomato by RNAi-mediated silencing of three
homologs of 1-aminopropane-1-carboxylate synthase gene Journal of Plant Physiology 170 (2013) 987ndash 995
Regulation of Ethylene Production
b Suppression of ACC oxidase gene expression
It catalyzes the oxidation of ACC to ethylene
The last step in the ethylene biosynthetic pathway
Down regulation through anti-sense technology
Hamilton et al 1990
Ye et al 1996
Xiong et al 2003
Ripening in papaya fruit is altered by ACC
oxidase cosuppression
Fig1Map of the construct pKYCPACOO-1 containing the ACC oxidase
fragment cloned in PKYLX80 in the sense orientation The ACC oxidase
fragment is flanked by the CaMV 35S promoter and the RUBISCO
terminator
Fig2 Ethylene production in papaya transgenic fruits
Rodolfo Loacutepez-Goacutemezet alTransgenic Res 1889ndash97 2009
c Insertion of the ACC deaminase gene
Regulation of Ethylene Production
The gene is obtained from Pseudomonas chlororaphis
(a common nonpathogenic soil bacterium)
It converts ACC to a different compound
Reduce the amount of ACC available for ethylene production
90-97 reduced ethylene production
Klee et al1991
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Antisense Technology
httpagbiosafetyunleduflashantisenseswf Journal of Plant Physiology170987ndash 9952013
Null Mutation of the MdACS3 Gene
Apple cultivars homozygous or heterozygous for null allelotype
showed no or very low expression of ripening-related genes and
maintained fruit firmness
Aide Wang 2009
RNAi-mediated silencing
Chimeric RNAi-ACS construct designed to target ACS
homologs
Delayed ripening and extended shelf life for sim45 days
Aarti Gupta Ram Krishna Pal Delayed ripening and improved fruit processing quality in tomato by RNAi-mediated silencing of three
homologs of 1-aminopropane-1-carboxylate synthase gene Journal of Plant Physiology 170 (2013) 987ndash 995
Regulation of Ethylene Production
b Suppression of ACC oxidase gene expression
It catalyzes the oxidation of ACC to ethylene
The last step in the ethylene biosynthetic pathway
Down regulation through anti-sense technology
Hamilton et al 1990
Ye et al 1996
Xiong et al 2003
Ripening in papaya fruit is altered by ACC
oxidase cosuppression
Fig1Map of the construct pKYCPACOO-1 containing the ACC oxidase
fragment cloned in PKYLX80 in the sense orientation The ACC oxidase
fragment is flanked by the CaMV 35S promoter and the RUBISCO
terminator
Fig2 Ethylene production in papaya transgenic fruits
Rodolfo Loacutepez-Goacutemezet alTransgenic Res 1889ndash97 2009
c Insertion of the ACC deaminase gene
Regulation of Ethylene Production
The gene is obtained from Pseudomonas chlororaphis
(a common nonpathogenic soil bacterium)
It converts ACC to a different compound
Reduce the amount of ACC available for ethylene production
90-97 reduced ethylene production
Klee et al1991
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Null Mutation of the MdACS3 Gene
Apple cultivars homozygous or heterozygous for null allelotype
showed no or very low expression of ripening-related genes and
maintained fruit firmness
Aide Wang 2009
RNAi-mediated silencing
Chimeric RNAi-ACS construct designed to target ACS
homologs
Delayed ripening and extended shelf life for sim45 days
Aarti Gupta Ram Krishna Pal Delayed ripening and improved fruit processing quality in tomato by RNAi-mediated silencing of three
homologs of 1-aminopropane-1-carboxylate synthase gene Journal of Plant Physiology 170 (2013) 987ndash 995
Regulation of Ethylene Production
b Suppression of ACC oxidase gene expression
It catalyzes the oxidation of ACC to ethylene
The last step in the ethylene biosynthetic pathway
Down regulation through anti-sense technology
Hamilton et al 1990
Ye et al 1996
Xiong et al 2003
Ripening in papaya fruit is altered by ACC
oxidase cosuppression
Fig1Map of the construct pKYCPACOO-1 containing the ACC oxidase
fragment cloned in PKYLX80 in the sense orientation The ACC oxidase
fragment is flanked by the CaMV 35S promoter and the RUBISCO
terminator
Fig2 Ethylene production in papaya transgenic fruits
Rodolfo Loacutepez-Goacutemezet alTransgenic Res 1889ndash97 2009
c Insertion of the ACC deaminase gene
Regulation of Ethylene Production
The gene is obtained from Pseudomonas chlororaphis
(a common nonpathogenic soil bacterium)
It converts ACC to a different compound
Reduce the amount of ACC available for ethylene production
90-97 reduced ethylene production
Klee et al1991
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
RNAi-mediated silencing
Chimeric RNAi-ACS construct designed to target ACS
homologs
Delayed ripening and extended shelf life for sim45 days
Aarti Gupta Ram Krishna Pal Delayed ripening and improved fruit processing quality in tomato by RNAi-mediated silencing of three
homologs of 1-aminopropane-1-carboxylate synthase gene Journal of Plant Physiology 170 (2013) 987ndash 995
Regulation of Ethylene Production
b Suppression of ACC oxidase gene expression
It catalyzes the oxidation of ACC to ethylene
The last step in the ethylene biosynthetic pathway
Down regulation through anti-sense technology
Hamilton et al 1990
Ye et al 1996
Xiong et al 2003
Ripening in papaya fruit is altered by ACC
oxidase cosuppression
Fig1Map of the construct pKYCPACOO-1 containing the ACC oxidase
fragment cloned in PKYLX80 in the sense orientation The ACC oxidase
fragment is flanked by the CaMV 35S promoter and the RUBISCO
terminator
Fig2 Ethylene production in papaya transgenic fruits
Rodolfo Loacutepez-Goacutemezet alTransgenic Res 1889ndash97 2009
c Insertion of the ACC deaminase gene
Regulation of Ethylene Production
The gene is obtained from Pseudomonas chlororaphis
(a common nonpathogenic soil bacterium)
It converts ACC to a different compound
Reduce the amount of ACC available for ethylene production
90-97 reduced ethylene production
Klee et al1991
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Regulation of Ethylene Production
b Suppression of ACC oxidase gene expression
It catalyzes the oxidation of ACC to ethylene
The last step in the ethylene biosynthetic pathway
Down regulation through anti-sense technology
Hamilton et al 1990
Ye et al 1996
Xiong et al 2003
Ripening in papaya fruit is altered by ACC
oxidase cosuppression
Fig1Map of the construct pKYCPACOO-1 containing the ACC oxidase
fragment cloned in PKYLX80 in the sense orientation The ACC oxidase
fragment is flanked by the CaMV 35S promoter and the RUBISCO
terminator
Fig2 Ethylene production in papaya transgenic fruits
Rodolfo Loacutepez-Goacutemezet alTransgenic Res 1889ndash97 2009
c Insertion of the ACC deaminase gene
Regulation of Ethylene Production
The gene is obtained from Pseudomonas chlororaphis
(a common nonpathogenic soil bacterium)
It converts ACC to a different compound
Reduce the amount of ACC available for ethylene production
90-97 reduced ethylene production
Klee et al1991
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Ripening in papaya fruit is altered by ACC
oxidase cosuppression
Fig1Map of the construct pKYCPACOO-1 containing the ACC oxidase
fragment cloned in PKYLX80 in the sense orientation The ACC oxidase
fragment is flanked by the CaMV 35S promoter and the RUBISCO
terminator
Fig2 Ethylene production in papaya transgenic fruits
Rodolfo Loacutepez-Goacutemezet alTransgenic Res 1889ndash97 2009
c Insertion of the ACC deaminase gene
Regulation of Ethylene Production
The gene is obtained from Pseudomonas chlororaphis
(a common nonpathogenic soil bacterium)
It converts ACC to a different compound
Reduce the amount of ACC available for ethylene production
90-97 reduced ethylene production
Klee et al1991
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
c Insertion of the ACC deaminase gene
Regulation of Ethylene Production
The gene is obtained from Pseudomonas chlororaphis
(a common nonpathogenic soil bacterium)
It converts ACC to a different compound
Reduce the amount of ACC available for ethylene production
90-97 reduced ethylene production
Klee et al1991
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Plants transformed with ACC
deaminase
No differences in softness
Major difference in degradation
of fruit that occurs following
ripening
Klee et al Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon
esculentum) Plants with Reduced Ethylene Synthesis Plant Physiol Vol 102 1993
Regulation of Ethylene Production
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Regulation of Ethylene Production
d Insertion of the SAM hydrolase gene
The gene is obtained from E coli T3 bacteriophage
SAM is converted to homoserine
The amount of its precursor metabolite is reduced
Matto 2002
Good et al 1994
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Regulation of Ethylene Production
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Regulation of Cell wall degradation
aPolygalacturonase (PG)
degrades pectin
Antisense RNA techniques
The transgenic fruit with decreased levels of PG activity
1)Do not get overly soft when ripe
2)Show less damage due to fungal infection and
3)Have elevated levels of soluble solids
Bird et al 1988
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Chimaeric polygalacturonase (PG)
gene
Produce a truncated PG transcript
constitutively
Expression of the endogenous PG
gene was inhibited
Regulation of Cell wall degradation
CJS Smith et al Expression of a truncated tomato polygalacturonase gene inhibits
expression of the endogenous gene in transgenic plants Mol Gen Genet224477-481
1999
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
bPectin methylesterase (PME)
Involved in metabolism of pectin
Break large polymers into shorter molecules
Antisense RNA approach
Transgenic fruit resulted in reduced pectin depolymerization
However there was no effect on firmness during ripening
Tieman et al1992
Hall et al1993
Regulation of Cell wall degradation
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
cβ-galactosidaseNormally upregulated during the early stages of ripening
Serves to remove pectic galactan side chains
Antisense regulation
dPhospholipase DHydrolyze phospholipids
An antisense phospholipase D (PLD) cDNA Construct
resulted in a 30-40 reduction of PLD activity in ripe fruits
Transgenic fruits were firmer possessed better red colour and flavour
Pinhero et al 2003
eDeoxyhypusine synthaseAntisense gen copy of Senescence-induced deoxyhypusine synthase and
senescence-induced elf-5a
Pleiotropic effects on growth and development of tomato
Transgenics ripened normally but exhibited delayed postharvest softening
Wang et al 2005
Regulation of Cell wall degradation
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Control of Ethylene Perception and signaling
Modifying ethylene receptors
The gene ETR1 encode an ethylene binding protein
Modified ETR1 lack the ability to respond to ethylene
Down-regulate specific tomato ethylene receptor isoforms using antisense
suppression have been reported for SlETR1 NR and SlETR4
Reporter genes related to ethylene responses and fruit ripening LeCTR1 and
SlEILs genes were also successfully silenced
Fu et al 2005
Zhu et al 2006
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Control of Ethylene Perception and signaling
Lucille Alexander Journal of Experimental Botany Vol 53 No 377JC Stearns BR Glick Biotechnology Advances 21 (2003) 193ndash210
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Anthocyanins Double the Shelf Life of
Tomatoes by Delaying Over ripening
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Binding of specific trans-acting factors to the cognate cis-elements
Governs the spatial and temporal expression of a number of inducible genes
Tomato
E8 (Deikman et al 1998)
2A11 (Vanand Houck 1993)
Apple
ACO (Atkinson et al1998)
Melon
cucumisin (Yamagata et al 2002)
WSP (Wu et al 2003)
Strawberry
GalUR (Agius et al2005)
Grape
VvAlb1 (Li and Gray 2005)
Banana
MaExp1 (Trivedi and Nath 2004)
Fruit specific and ripening related
promoterscis-elements
Research in Environment and Life Sciences 2008
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Advantages of Delayed fruit ripening
Assurance of top quality
Allowing the fruits to exude full quality
Consumers will get value for their money
Widening of market opportunities
Reduction in postharvest losses
httpwwwisaaaorgkc
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
References
James J Giovannoni Genetic Regulation of Fruit Development and Ripening The
Plant Cell Vol 16 S170ndashS180 2004
Antonio J Matas et al Biology and genetic engineering of fruit maturation for enhanced
quality and shelf-life Current Opinion in Biotechnology 20197ndash203 2009
V Prasanna et al Fruit Ripening PhenomenandashAn Overview Critical Reviews in Food
Science and Nutrition 471ndash19 2007
M Bouzayen et al Mechanism of Fruit Ripening Open Archive TOULOUSE Archive
Ouverte Eprints ID 4525
Aide Wang et al Null Mutation of the MdACS3 Gene Coding for a Ripening-Specific
1-Aminocyclopropane-1-Carboxylate Synthase Leads to Long Shelf Life in Apple Fruit
Plant Physiology Vol 151 pp 391ndash399 2009
Rodolfo Loacutepez-Goacutemez et al Ripening in papaya fruit is altered by ACC oxidase
Cosuppression Transgenic Res 1889ndash97 2009
Aarti Gupta et al Delayed ripening and improved fruit processing quality in tomato by
RNAi-mediated silencing of three homologs of 1-aminopropane-1-carboxylate
synthase gene Journal of Plant Physiology 170987ndash 995 2013
Liu C et al Cloning of 1-aminocyclopropane-1-carboxylate (ACC) synthetase cDNA
and the inhibition of fruit ripening by its antisense RNA in transgenic tomato plants Chin
J Biotechnol 199814(2)75-84
Gray J et al Molecular biology of fruit ripening and its manipulation
with antisense genes Plant Mol Biol 1992 May19(1)69-87
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
Oeller PW et al Reversible inhibition of tomato fruit senescence by antisense RNA
Science 1991 Oct 18254(5030)437-9
Harpster MH Constitutive overexpression of a ripening-related pepper endo-14-beta-
glucanase in transgenic tomato fruit does not increase xyloglucan depolymerization or
fruit softening Plant Mol Biol 2002 Oct50(3)357-69
Brummell DA et al Cell wall metabolism in fruit softening and quality and its
manipulation in transgenic plants Plant Mol Biol 2001 Sep47(1-2)311-40
Websiteshttpagbiosafetyunleduflashantisenseswf
httpwwwisaaaorgkc
httpwwwukessayscom essaysbiologyquality-and-shelf-life-of-fruits-and-
vegetablesphp
httpshodhgangainflibnetacinbitstream1060340711616_referencespdf
BooksBiology and biotechnology of the plant hormone ethylene
Edited by- A Khanellis
Transgenic plants and crops
Edited by- M Dekkerlne
References
THANK YOU
THANK YOU
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