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    DroughtResearch

    GroupPhD research areas

    2012

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    The teams

    Forward genetics: from phenotype to geneWe observe and analyse the characteristics of drought tolerance in wheat and barley

    and seek to identify the genes responsible of the tolerance. The following projects have

    been initiated:

    Detailed physiological analysis ofdrought tolerant and intolerant wheat

    lines under controlled environment

    and field conditions

    Genetic analysis of drought toleranceunder field and growth room

    conditions in three large populations

    of wheat Genetic dissection of root

    development and architecture under

    normal and drought conditions

    Development of metabolite andtranscript profiles of parental lines

    under water limited conditions

    Reverse genetics: from gene to phenotypeWe seek to isolate gene sequences

    important for conferring drought

    tolerance from both model and crop

    species. Cloned gene sequences are

    introduced into our target crop

    species by either biolistics (wheat) or

    Agrobacterium-mediated

    transformation (barley). Transgenic

    plants are then assayed under

    controlled and field conditions fordrought tolerance. Project areas

    include:

    Bioinformatics and the identificationof drought related gene sequences

    Transcription factors and theregulation of drought-stress responses

    Development of commercially viabledrought tolerant GM wheat and barley

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    Our collaboratorsBBG University of Adelaide (South Australia)

    DPI La Trobe University (Victoria)

    INRA Clermont-Ferrand (France)

    SCRI Dundee (Scotland)

    IPK Gatersleben (Germany)

    CIMMYT (Mexico)

    Pioneer Hi-Bred International/DuPont (USA)

    INRA Montpellier (France)

    University of Bologna (Italy)

    The techniques

    Molecular biology

    DNA and RNA extractions

    PCRs and restriction digests

    Transcript profiling by microarrays and

    sequencing

    Metabolite and protein profiling

    Confocal microscopy

    BAC library screening

    BiotechnologyCloning

    Biolistics andAgrobacterium-mediated plant

    transformation

    Fluorescent reporters (GFP, mCherry)

    Bioinformatics

    Plant Physiology

    High throughput phenotyping by imaging

    Chlorophyll content

    Stomatal conductance

    Photosynthesis

    Plant and soil water potentials

    Canopy temperature

    Root and shoot anatomy

    Root morphology

    Genetics

    QTL mapping

    Molecular markers

    Positional cloning

    Marker assisted selection

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    Potential PhD supervisors:

    Dr Ute Baumann

    Dr Omid Eini

    Dr Delphine Fleury

    Dr Chunyuan Huang

    Dr Nataliya Kovalchuk

    Prof. Peter Langridge

    Dr Sergiy Lopato

    Prof Diane Mather

    Dr Boris Parent

    Dr Bujun Shi

    Dr Ryan Whitford

    Adelaide node of the ACPFG at the Plant Genomics Centre

    (Waite Campus, University of Adelaide)

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    PhD projects:

    Project title: Genetic and physiological characterization of tolerance to heat-

    induced floret sterility in wheat

    SUPERVISOR: Dr Nick Collins

    CONTACTDETAILS:[email protected]

    BACKGROUND

    Brief periods of heat stress severely impact on wheat production, and this situation will worsen with

    climate change. However, it is difficult for breeders to select for heat tolerance in the field, because

    heat affects wheat in different ways depending on the growth stage, and because natural heat

    events are unpredictable in severity and timing. Therefore, there is an urgent need for reproducible

    growth chamber assays for heat tolerance that are relevant to the field, molecular markers linked to

    genes controlling heat tolerance, or cloned heat tolerance genes for transformation breeding.

    One type of heat damage is the floret sterility (decrease grain set) following heat events at around or

    just before pollination. Anecdotal reports indicate that Australian durum wheat varieties are more

    prone to this form of heat damage than bread wheats, however this difference needs to be formally

    tested. Besides this, there is virtually nothing known about how much wheat genotypes naturally

    vary for tolerance to heat-induced sterility effects.

    AIMS

    Use a growth chamber to characterize the precise developmental stages where durum andbread wheat are most sensitive to heat-induced sterility. Use this information to design

    tolerance assays targeting sterility effects of heat.

    Screen local and exotic durum and bread wheat varieties for variation in tolerance to heat-induced sterility.

    Characterize the biological basis for the tolerance in various sources. Use new or established populations to map chromosome regions (QTLs) controlling variation

    in tolerance.

    Initiate positional cloning of heat tolerance genes via candidate genes.

    TECHNIQUES TO BE USED

    Growth chamber assays for heat tolerance. Characterize tiller stages by examining developing spikes by microscopy (stage of meiosis or

    development of female reproductive structures).

    In vitro pollen viability/tube-growth assays. Chlorophyll fluorescence measurements (Fv/Fm). SSR, DArT and SNP markers. Comparative mapping and sequence analysis for targeted marker generation and candidate

    gene identification.

    Gene transcript quantification and tissue-localization by qRT-PCR.

    mailto:[email protected]:[email protected]:[email protected]:[email protected]
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    REFERENCES

    Barnabas B, Jager K, Feher A (2008). The effect of drought and heat stress on reproductive processes

    in cereals. Plant, Cell and Environment 31:11-38.

    Saini HS, Sedgley M, Aspinall D (1984). Developmental anatomy in wheat of male sterility induced byheat stress, water deficit or abscisic acid. Australian Journal of Plant Physiology 11:243-253.

    Singh SK, Kakani VG, Brand D, Baldwin B, Reddy KR (2008). Assessment of cold and heat tolerance of

    winter-grown canola (Brassica napus L.) cultivars by pollen-based parameters. Journal of Agronomy

    & Crop Science 194:225-236.

    Heat treatment of

    wheat plants using a

    growth chamber. Plants

    are grown in thegreenhouse before and

    after a brief heat

    treatment applied at a

    specific developmental

    stage.

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    Project title: Positional cloning of QTL for drought tolerance in wheat

    SUPERVISORS:Dr Delphine Fleury, Prof. Peter Langridge

    CONTACT DETAILS:[email protected],[email protected]

    BACKGROUND

    The key objective of the drought program of ACPFG is to generate detailed knowledge of the

    mechanisms of drought adaptation under the Mediterranean type growing conditions, with a view

    to developing plants tolerant to multiple components: osmotic & oxidative stress, heat, dehydration.

    This type of drought is characterised by water deficit at the late stages of crop development, usually

    during flowering and grain filling.

    We have identified several QTLs of wheat controlling yield in dry environment. Four QTL are

    targeted for map-based cloning, for which we already have large populations of recombinant inbred

    lines. The availability of new genomics resources, particularly the next-generation sequencing data,

    enables now to make tremendous progress in gene cloning in wheat. We are now increasing the

    number of markers for fine-mapping of each region to the resolution of each local map to

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    Project title: Genetic study of floral architecture for hybrid wheat systemSUPERVISORS:Dr Delphine Fleury, Dr Ryan Whitford

    CONTACT DETAILS:[email protected],[email protected]

    BACKGROUNDOne of the Green revolution technologies was the hybrid seeds. Hybrid plants obtained by inter-

    crossing inbred lines show an increase in biomass and production. This heterotic effect is particularly

    strong in out-breeding species such as maize. Past studies showed that yield increase is possible to

    achieve in hybrid wheat compared to the conventional inbred lines. However due to its self-

    pollinated nature, inter-crossing plants is difficult. One of the factors that impair out-crossing in

    wheat is its flower architecture and biology: the spike is compact, male and female are enclosed in

    spikelet, anthers and styles are short, flowering time isnt synchronised between male and female

    parental lines.

    Chasmogamic species are characterized by open flowers and exposed stamens and styles that

    facilitate inter-pollination. These traits are usually controlled by few major genes. Heritability of

    flower architecture is medium to high suggesting that progress could be made in improving thecross-pollinating ability of parental lines. The aim of this project is to identify wheat loci and genes

    that will increase chasmogamy and facilitate inter-crossing using hybridization systems.

    AIMS AND SIGNIFICANCE:Identify QTL and genes controlling flower architecture of wheat for increasing

    hybridization rate in hybrid seed production.

    TECHNIQUES TO BE USED:

    Molecular markers, phenotyping of floral development, statistical analysis, genome sequence

    analysis, genetic mapping and QTL software

    REFERENCESTester M. and Langridge P. 2010. Breeding technologies to increase crop production in a changing

    world. Science 327: 818-822.

    Jordaan JP. Hybrid wheat: advances and challenges. 1996. In: MP Reynolds, S Rajaram and A McNab

    (eds) Increasing yield potential in wheat: breaking the barriers, Mexico, CIMMYT, pp 66-75.

    mailto:[email protected]:[email protected]:[email protected]:[email protected]:[email protected]
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    Project title: QTL mapping of root traits associated with efficient wheat root

    systems in drought environments

    SUPERVISORS:Dr Chunyuan Huang, Prof. Peter Langridge

    CONTACT DETAILS:[email protected], [email protected]

    BACKGROUND

    Root systems determine the ability of a plant to capture available water and nutrients. Most of past

    research had concentrated on above-ground characteristics of plants, while below-ground

    characteristics were mostly neglected. Root proliferation is sensitive to environmental conditions.

    The current techniques for root phenotyping are labour-intensive and error-prone, especially for

    field-grown plants. We have developed a novel root phenoptyping technique in collaboration with

    researchers in SARDI, using quantitative real time PCR. This new technique provides accurate

    measurements of living root cells, and therefore, it improves efficiency and accuracy of root

    phenotyping. The new root technique has been trialled for high-throughput root phenotyping of

    field-grown cultivars. Our results show that restrained root growth is important for high grain yield

    in Mediterranean drought environments. The QTL mapping of restrained root systems will open

    opportunities for genetic improvement of crop tolerance to drought.

    AIMS AND SIGNIFICANCE:Identifying QTLs associated with efficient wheat root systems in drought

    environments, and improving wheat tolerance to drought

    TECHNIQUES TO BE USED:

    Measurement of root traits using a range of techniques(WinRHIZO), Q-PCR, genetic mapping and

    QTL software, DNA sequencing and gene analysis

    REFERENCESGenc Y, Huang CY, Langridge P (2007) A study of the role of root morphological traits in growth of

    barley in zinc-deficient soil. J. Exp. Bot. 58, 2775-2784

    McKay A, Riley IT, Hartley D, Wiebkin S, Herdina, Li G, Coventry S, Hall S, Huang CY (2008) Studying

    root development in soil using DNA technology: idea to impact.(http://www.regional.org.au/au/asa/2008/plenary/biotechnology/5945_mckay.htm#TopOfPage)

    Root analysis software (WinRhizo)

    mailto:[email protected]:[email protected]:[email protected]:[email protected]:[email protected]://www.regional.org.au/au/asa/2008/plenary/biotechnology/5945_mckay.htm#TopOfPagehttp://www.regional.org.au/au/asa/2008/plenary/biotechnology/5945_mckay.htm#TopOfPagehttp://www.regional.org.au/au/asa/2008/plenary/biotechnology/5945_mckay.htm#TopOfPagehttp://www.regional.org.au/au/asa/2008/plenary/biotechnology/5945_mckay.htm#TopOfPagemailto:[email protected]:[email protected]
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    Transgenic barley roots express GFP, indicating new living roots in green (red arrow) and the dead

    roots (white arrow).

    Field trials for root phenotyping using DNA technique

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    Project title: Analysis of drought inducible promoters from wheat

    SUPERVISORS:Dr Nataliya Kovalchuk, Dr Omid Eini, Dr Sergiy Lopato

    CONTACT DETAILS:

    [email protected],[email protected], [email protected]

    BACKGROUND:

    Drought is among the major environmental factors limiting crop productivity worldwide. An

    important response to this stress is the temporal and spatial modulation of transcription of specific

    sets of genes, which enables the plant to rapidly adapt to altered environmental conditions. The

    transcriptional response of plants to drought stress is controlled by numerous transcription factors

    (Agarwal et al., 2006; Shinozaki et al., 2007). We have successfully used wheat and barley cDNA

    libraries for a yeast one-hybrid (Y1H) screen for cDNAs encoding transcription factors (TFs) up-

    regulated by drought (Lopato et al., 2006). Some TFs have been over-expressed in transgenic barley

    and wheat under constitutive promoters and generated transgenic plants demonstrate increased

    drought tolerance. Unfortunately, expression of TFs under constitutive promoters often leads to

    undesirable developmental phenotypes (Morran et al., in press). To overcome this problem we

    prepared a collection of drought inducible and tissue-specific promoters, which we are currently

    testing in transgenic wheat and barley. Several wheat promoters will be analysed for spatial and

    temporal expression in transgenic wheat and/or barley in the absence of stress and then under

    drought and several other abiotic stresses (Li et al., 2008; Rai et al., 2009; Kovalchuk et al., 2010).

    Mapping of stress specific cis-elements in some of the promoters and isolation of up-stream

    transcription factors using Y1H system will be inclusive to this project.

    AIM OF THE PROJECT:

    To characterise and test different types of drought inducible promoters from wheat using transient

    and stable expression of promoter-reporter gene constructs in wheat and barley.

    TECHNIQUES TO BE USED

    1. Analysis of promoter structure, identification of stress responsive cis-elements, and isolationof upstream transcription regulators using the Y1H screen.

    2. Careful analysis of transgenic wheat and barley development. These transgenics expressdrought tolerance TF under several different drought inducible and spike specific promoters.

    3. Compare transgenic wheat and barley exhibiting differing expression patterns of the samedrought tolerance TF for survival and yield under drought stress.

    4. Participation in field trials.REFERENCES:

    Agarwal PK et al. Role of DREB transcription factors in abiotic and biotic stress tolerance in plants.

    Plant Cell Rep. 2006 25(12):1263-1274.

    Kovalchuk N et al. Defensin promoters as potential tools for engineering disease resistance in cereal

    grains. Plant Biotechnol J. 2010 8(1):47-64

    Li M et al. Spatial and temporal expression of endosperm transfer cell-specific promoters in

    transgenic rice and barley. Plant Biotechnol J. (2008) 6(5):465-76.

    Lopato S et al., Isolation of plant transcription factors using a modified yeast one-hybrid system.

    Plant Methods 2006, 2:3-17

    Rai M et al., Comparative functional analysis of three abiotic stress-inducible promoters in transgenic

    rice. Transgenic Res. 2009 18(5):787-99.

    Shinozaki K and Yamaguchi-Shinozaki K. Gene networks involved in drought stress response and

    toleranceJ. Exp. Bot. 2007 58:221-7.

    mailto:[email protected]:[email protected]:[email protected]:[email protected]:[email protected]:[email protected]:[email protected]:[email protected]:[email protected]:[email protected]
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    Fig. 1. Trangenic wheat lines with drought inducible over-expression of DREB/CBF demonstrate

    improved survival under stringent drought conditions

    control Line BW8-10Line BW8-6Line BW8-2

    14 daysno water

    7 daysafter re-watering

    14 daysafter re-watering

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    Project title: Isolation and characterization of genes involved in the

    formation and regulation of drought responsive NF-Y transcription

    complex(es)

    SUPERVISORS: Dr Sergiy Lopato, Dr Omid Eini, Dr Maria Hrmova, Prof. Peter Langridge

    CONTACT DETAILS:[email protected] ,[email protected]

    BACKGROUND:

    Drought stress is a major constraint to the production and yield stability of wheat in Australia. Plants

    respond to drought by the temporal and spatial transcriptional modulation genes - a response

    orchestrated by various transcription factors (Agarwal et al., 2006; Shinozaki et al., 2007). Nuclear

    Factor Y (NF-Y) is a trimeric complex that binds to the CCAAT box of plant promoters, with each

    subunit being required for DNA binding (Fig. 1). At least 37 genes for NF-Y subunits have been

    identified in wheat, and at least some of these are up-regulated by drought (Stephenson et al.,

    2007). Furthermore, over-expression of NF-YB and NF-YA genes in transgenic Arabidopsis and maizeplants can confer drought tolerance without negatively affecting plant development (Nelson et al.,

    2007; Li et al, 2008). However, it is unclear which of these subunits participate in particular

    complexes, and whether subunits in a complex can be substituted without affecting functionality.

    We have used wheat and maize spike and developing grain subjected to drought stress to make

    cDNA libraries for Y2H screens (Lopato et al., 2006). We are now screening these libraries to identify

    interacting partners of NF-Y subunits, which may include NF-Y subunits or other types of factors that

    cooperate with NF-Y trimeric complexes during drought stress (unpublished data). Some of the TFs

    identified have already been over-expressed in transgenic barley and wheat under constitutive and

    inducible promoters.

    AIM OF THE PROJECT:To isolate wheat genes encoding drought related members of the NF-Y complex using PCR-based

    cloning and a yeast two-hybrid (Y2H) screen, and to characterise transgenic wheat and barley plants

    with constitutive and drought inducible over-expression of two NF-Y subunits.

    TECHNIQUES TO BE USED

    The project will involve further Y2H screens for NF-Y interacting factors, characterisation of these

    genes, and preparation of constructs for wheat and barley transformation. Two transgenics over-

    expressing NF-Y genes are already available for drought tolerance evaluation. The project may also

    involve structural modelling and experimental verification of protein-protein and protein-DNA

    interactions involving NF-Y trimeric complexes.

    Specific techniques will include: Isolation of genes relevant to NF-Y complexes using PCR-based cloning and the Y2H screen of

    cDNA libraries prepared from spike, developing grain and root subjected to drought stress

    Analysis of expression of isolated genes using quantitative PCR (Q-PCR). Modelling of NF-Y protein-protein and protein-DNA interactions (optional) Analysis of transgenic wheat and barley plants with constitutive and inducible overexpression of

    two NF-Y factors for survival and performance under drought stress.

    Participation in field trials.REFERENCES:

    Agarwal PK et al. Role of DREB transcription factors in abiotic and biotic stress tolerance in plants.

    Plant Cell Rep. 2006 25(12):1263-1274

    mailto:[email protected]:[email protected]:[email protected]:[email protected]:[email protected]:[email protected]:[email protected]:[email protected]
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    Li WX, Oono Y, Zhu J, He XJ, Wu JM, Iida K, Lu XY, Cui X, Jin H, Zhu JK.The Arabidopsis NFYA5

    transcription factor is regulated transcriptionally and posttranscriptionally to promote drought

    resistance. Plant Cell. 2008 Aug;20(8):2238-51.

    Lopato S, Borisjuk L, Milligan AS, Shirley N, Bazanova N, Parsley K, Langridge P.

    Systematic identification of factors involved in post-transcriptional processes in wheat grain. Plant

    Mol Biol. 2006 62(4-5):637-53Nelson DE, Repetti PP, Adams TR, Creelman RA, Wu J, Warner DC, Anstrom DC, Bensen RJ, Castiglioni

    PP, Donnarummo MG, Hinchey BS, Kumimoto RW, Maszle DR, Canales RD, Krolikowski KA, Dotson

    SB, Gutterson N, Ratcliffe OJ, Heard JE. Plant nuclear factor Y (NF-Y) B subunits confer drought

    tolerance and lead to improved corn yields on water-limited acres. Proc Natl Acad Sci U S A. 2007

    104(42):16450-5.

    Shinozaki K and Yamaguchi-Shinozaki K. Gene networks involved in drought stress response and

    toleranceJ. Exp. Bot. 2007 58:221-7

    Stephenson TJ, McIntyre CL, Collet C, Xue GP. Genome-wide identification and expression analysis of

    the NF-Y family of transcription factors in Triticum aestivum. Plant Mol Biol. 2007 65(1-2):77-92.

    Fig. 1. HAP (Heme Activator Protein) Complex or CBF (CCAAT Binding Factor) or

    NF-Y (Nuclear Factor Y) Complex

    C C A A T

    C C A A T

    HAP3/

    CBFA/

    NF-YB

    HAP5/

    CBFC/

    NF-YC

    HAP2/

    CBFB/

    NF-YA HAP5/

    CBFC/

    NF-YC

    HAP3/

    CBFA/

    NF-YB

    HAP2/

    CBFB/

    NF-YA

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    Project title: Isolation and characterization of root specific and drought

    inducible transcription factors from wheat roots subjected to drought and

    salt stress

    SUPERVISORS: Dr Sergiy Lopato, Dr Chunyuan Huang, Prof. Peter Langridge

    CONTACT DETAILS:[email protected] ,[email protected]

    BACKGROUND:

    Drought is a major constraint to the production and yield stability of wheat in Australia. Root

    characteristics, especially root length, root length density, and the number of thick roots, are critical

    for the exploitation of available soil water and for healthy above-ground growth. An important

    response to drought is the temporal and spatial transcriptional modulation of specific genes, which

    enables the plant roots to adapt to altered environmental conditions. The transcriptional response

    of plants to drought stress is controlled by various transcription factors (Agarwal et al., 2006;

    Shinozaki et al., 2007). However, very little is known about the transcription factors that control rootgrowth, particularly in a drought specific manner (Hirsch and Oldroyd, 2009;Coudert et al., 2010).

    We have made cDNA libraries from wheat and barley spike and developing grain subjected to

    drought stress and successfully used these in Y1H (Fig. 1) and Y2H screens for transcription factors

    (TFs) up-regulated by drought (Lopato et al., 2006a, 2006b). Over-expression of some of these TFs in

    transgenic barley and wheat under constitutive and inducible promoters was found to provide

    improved survival under severe drought conditions (Morran et al., in press). We have also made

    several cDNA libraries from wheat and maize roots under drought and salt stress, and initial tests

    suggest that these libraries provide a good source of root specific and stress inducible TF genes.

    AIM OF THE PROJECT:

    To isolate members of several families of root specific and drought inducible transcription factorsusing yeast one-hybrid (Y1H) or two-hybrid (Y2H) screens, together with PCR-based cloning, and to

    characterise these genes for their spatial, temporal and stress inducible patterns of expression.

    TECHNIQUES TO BE USED

    The project will include isolation of TFs from root libraries using Y1H and Y2H screens, as well as PCR-

    based cloning, and assessment of TF expression under well watered or drought conditions. Cloning

    of root specific genes and promoters and mapping of stress specific and root specific cis-elements in

    selected promoters will be part of this project.

    Specific activities include:

    Isolation of several transcription factors using Y1H and Y2H screens of root cDNA libraries andPCR-based cloning

    Analysis of expression of isolated TFs using quantitative PCR (Q-PCR) on RNA prepared fromstressed plants.

    Isolation of full length genes and promoters for root specific TFs. Analysis of promotersequences.

    Preparation of constructs for transformation of wheat and barleyREFERENCES:

    Agarwal PK et al. Role of DREB transcription factors in abiotic and biotic stress tolerance in plants.

    Plant Cell Rep. 2006 25(12):1263-1274

    Coudert Y, Prin C, Courtois B, Khong NG, Gantet P. Genetic control of root development in rice, the

    model cereal. Trends Plant Sci. 2010 15(4):219-26

    mailto:[email protected]:[email protected]:[email protected]:[email protected]:[email protected]:[email protected]:[email protected]:[email protected]
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    Hirsch S, Oldroyd GE. GRAS-domain transcription factors that regulate plant development. Plant

    Signal Behav. 2009 4(8):698-700

    Lopato S et al., Isolation of plant transcription factors using a modified yeast one-hybrid system.

    Plant Methods 2006, 2:3-17

    Lopato S, Borisjuk L, Milligan AS, Shirley N, Bazanova N, Parsley K, Langridge P.

    Systematic identification of factors involved in post-transcriptional processes in wheat grain. PlantMol Biol. 2006 62(4-5):637-53

    Shinozaki K and Yamaguchi-Shinozaki K. Gene networks involved in drought stress response and

    toleranceJ. Exp. Bot. 2007 58:221-7

    Fig. 1. Schematic representation of the selection of positive clones during Y1H screen.

    Isolation of TFs using Y1H screen

    Bait sequence TATA HIS3selection gene

    Transcription if hybrid

    protein interacts with

    bait sequenceADLibrary protein

    Selection for positive

    growth on CM HisLeu

    3-AT and X-GAL plates

    GAL4 TATA MEL1 selection gene

    ADNon-specific protein

    Transcription if non-specific

    protein interacts with

    bait sequence

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    Project title: Determination of microRNAs involved in drought tolerance in

    barley or wheat

    SUPERVISORS:Dr Bu-Jun Shi, Prof Peter Langridge

    CONTACT DETAILS:[email protected], [email protected]

    BACKGROUND

    The molecular biology of gene silencing represents one of the most important scientific advance in

    recent years. MicroRNAs (miRNAs) are one of the two key players in this process and have become a

    hot topic in molecular biology. miRNAs are a distinct class of small non-coding RNAs of 18-25

    nucleotides in length and conserved in eukaryotic organisms. In animals, miRNAs regulate 68% of

    total genes and participate in the regulation of almost every cellular process. In plants, miRNAs

    function very much like animal miRNAs, but play additional roles in development and adaptive

    responses to stresses. Wheat and barley are two most important and widely consumed cereals in

    the world. Their productivity, however, is severely limited by drought stress. To develop barley and

    wheat tolerant to drought stress, identifying key regulators of stress tolerance is crucial. Expression

    of some miRNAs is induced by drought, suggesting a role for miRNAs in drought stress. To

    investigate the role of miRNAs to drought stress in barley and wheat, we have constructed 2 small

    RNA libraries from barley and 6 small RNA libraries from wheat under normal and drought

    conditions. High throughput sequencing of these libraries reveals a number of candidate miRNAs,

    some of which are highly expressed, while others are lowly expressed under drought. The next step

    is to design experiments to confirm which miRNAs are truly associated with drought stress and

    investigate options for enhancing stress tolerance.

    AIMS AND SIGNIFICANCE:

    Determine miRNA function in drought stress responses, and express miRNA in transgenic barley or

    wheat to improve drought tolerance. The expected results would represent a significant advance in

    cereal miRNA research and the effort to enhance crop drought tolerance. To date, no barley miRNA

    has been identified, let alone their role in drought stress. Although 32 miRNAs from wheat have

    been reported, their function has not yet been experimentally confirmed.

    TECHNIQUES TO BE USED:

    Most of molecular biology techniques such as expression cloning, polymerase chain reaction (PCR),

    Southern blot hybridization, northern blot hybridization and western blotting. Other techniques

    include biochemical techniques, cell biology techniques, molecular genetic techniques and a range of

    bioinformatics approaches.

    REFERENCESBaulcombe D (2007). Amplified silencing. Science 315: 199200.

    Carrington J, Ambros V (2003). Role of microRNAs in plant and animal development. Science 301:

    3368.

    Khraiwesh B, Arif MA, Seumel GI, Ossowski S, Weigel D, Reski R, Frank W (2010). Transcriptional

    control of gene expression by microRNAs. Cell140:111-22.

    mailto:[email protected]://en.wikipedia.org/wiki/Southern_blothttp://en.wikipedia.org/wiki/Northern_blothttp://en.wikipedia.org/wiki/Northern_blothttp://en.wikipedia.org/wiki/Southern_blotmailto:[email protected]
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    The mechanism of Micro RNA generation and function

    Genomic DNA

    Pri-miRNA

    Pre-miRNA

    Transcription

    Enzymatic process

    Process further

    miRNA-miRNA* duplex (~21 nt)

    Unwind

    Single-stranded

    RISC assembly

    Bind to target mRNA

    Cleave target mRNA

    More miRNA(X)Less miRNA(X)

    Barley growth under drought

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    Scholarships

    ACPFG PhD scholarships consist of $25,000 tax free per year, plus support to attend an

    international conference and for professional development.

    Students applying for ACPFG PhD scholarships must be Australian citizens, permanentresidents, or New Zealand Citizens.

    International students can apply for these ACPFG scholarships only if they have received a

    university tuition fee waiver (or if they receive a stipend which covers the tuition fee).

    Further scholarships are also available through the:

    University of Adelaide

    http://www.adelaide.edu.au/graduatecentre/scholarships/postgrad/

    University of South Australia

    http://www.unisa.edu.au/scholarship/default.asp)University of Melbourne

    http://www.futurestudents.unimelb.edu.au/research/scholarships.html

    Contact

    Education manager: Dr Monica Ogierman

    Email:[email protected]: (08) 8303 6725

    Fax: (08) 8303 7102

    Australian Centre for Plant Functional

    Genomics (ACPFG)

    School of Agriculture, Food and Wine

    University of Adelaide Waite Campus

    Plant Genomics Centre

    Hartley Grove, Urrbrae

    Postal: PMB1 Glen Osmond SA 5064

    http://www.acpfg.com.au

    http://www.adelaide.edu.au/graduatecentre/scholarships/postgrad/http://www.adelaide.edu.au/graduatecentre/scholarships/postgrad/http://www.unisa.edu.au/scholarship/default.asphttp://www.unisa.edu.au/scholarship/default.asphttp://www.futurestudents.unimelb.edu.au/research/scholarships.htmlhttp://www.futurestudents.unimelb.edu.au/research/scholarships.htmlmailto:[email protected]:[email protected]:[email protected]://www.acpfg.com.au/http://www.acpfg.com.au/http://www.acpfg.com.au/mailto:[email protected]://www.futurestudents.unimelb.edu.au/research/scholarships.htmlhttp://www.unisa.edu.au/scholarship/default.asphttp://www.adelaide.edu.au/graduatecentre/scholarships/postgrad/
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    Additional reading

  • 7/31/2019 Drought_PhD Booklet 2012 v2

    22/22