Transgenesis, Intragenesis & Cisgenesis

I A B r i e f R e v i e w I

Overview

● Introduction● Conventional plant breeding● Transgenesis● Cisgenesis● Cisgenesis v/s Transgenesis● Cisgenesis v/s Conventional breeding● Intragenesis● Cisgenesis v/s Intragenesis● Cisgenesis, Intragenesis and Transgenesis● Transgenesis, Intragenesis, Cisgenesis and Conventional

breeding● Conclusion● References

Introduction

Problems related to agriculture & food security have led to the development of new technologies to curb those & meet the ever-increasing food demand.

In the past Conventional breeding

New technologiesPresently

Cisgenesis

Transgenesis

Intragenesis

Moreover, they hold the potential to produce crops with better, improved agricultural traits & of consumer acceptability.

Biotech

Conventional Plant BreedingPlant breeding is the science of improving genetic properties of cultivated crops. Conventional techniques were:

1. F1 hybrid breeding (hybridisation)2. Bridge crossing3. Repeated backcrossing

1 3

2

Conventional Plant Breeding

4. Cut Style technique

Other techniques such as combination breeding, mutation induction and anther/microspore culture were developed and are still in use.

5. Grafting on the style

TransgenesisGenetic modification by either the expression of a foreign gene or the suppression of an endogenous protein to modify a function. Genes/DNA can be moved between any species.

Examples:● First GMO commercialised (1994): Flavr Savr™ tomato

○ Longer shelf life (antisense RNA regulating the level of polygalacturonase enzyme involved in fruit ripening)

○ In 1996, soybean, canola, cotton and maize, herbicide- resistant transgenic lines were introduced to simplify weed-control practices.

(Espinoza et al., 2013)

Although it’s a promising tool for agriculture and enhancing economic development, major concern for public acceptance is an important factor.

CisgenesisSchouten et al. (2006) definition of ‘cisgenic plant’:“A crop plant that has been genetically modified with one or more genes (containing introns and flanking regions such as native promoter and terminator regions in a sense orientation) isolated from a crossable donor plant”.

That is:● It has all the necessary regulatory elements of a natural gene (a

perfect copy).

(Espinoza et al., 2013)

Examples (Telem et al., 2013): - Cisgenic apple which confer scab resistance- Cisgenic barley with improved phytase activity

Cisgenesis VS Transgenesis

● Cisgenesis: transfer of genes between crossable species or the plant itself○ same techniques as in traditional plant breeding can be

used, and as safe○ new traits without use of foreign genes○ no environmental risks

● Transgenesis: transfer between non-crossable species○ Recombination used○ Regulations concerning GMOs are based on transgenes

only: thus strict○ Foreign genes ( bacteria, sexually incompatible species)

(Hou et al., 2014)

Cisgenesis VS Transgenesis

Hence, there is a need to differentiate between the two to avoid problems like:● Delaying research & application of improved plant varieties.● Presently, many genes from wild varieties are being isolated

with potential cisgenetic application, but restriction due to laws.○ Except in Canada & Australia.

● European Food Safety Authority has done a safety assessment.○ Conclusions: same hazards as traditional breeding &

same risks in transfer technology.

(Hou et al., 2014)

Cisgenesis VS Traditional breeding● Cisgenics only contain gene(s) of interest unlike crops that

have been modified by classical breeding, which contain undesired genetic elements.

● Application of cisgenesis would produce results in only a few years of experimentation whereas traditional breeding would take decades to reach a cultivar with desirable traits.E.g: Breeding of apple for scab resistance took 50 years (Hou

et al., 2014).

Intragenesis

● Transfer of genes between crossable species.

● An alternative to transgenics, just like cisgenics.

● However, unlike cisgenes, intragenes are hybrid genes.○ i.e they can have genetic elements from different genes &

loci, thus○ by using different promoter or terminator regions,

expression of genes can be modified.○ Hence, there is the possibility of new gene

recombinations by in vitro rearrangements of functional genetic elements.

(Holme et al., 2013)

Cisgenesis VS Intragenesis

● Both more generally accepted by the public than the other techniques since○ they do not involve recombination between non-sexually

compatible organisms.● No foreign sequences present in the final organism.

○ e.g selection marker genes & vector-backbone sequences● However, cisgenesis is more restrictive than intragenesis.

Specific alleles/genes present in the breeders’ gene pool are introduced, without any change to the DNA sequence, into new varieties.

Genes can be designed using genetic elements from other plants with the same sexually compatibility gene pool.

Thus, the coding regions of a gene can be combined with promoters and terminators from different genes (from the same sexually compatibility gene pool).

IntragenesisCisgenesis

Cisgenesis can accelerate the breeding of species with long reproduction cycles since it does not contain the linkage drag.

Differences between Traditional breeding, Intra & Cisgenesis

Intragenesis Traditional breeding Cisgenesis

Regulatory elementsNew compositions of

coding sequences and promoters are made (7)

The gene has its native promoter, introns and

terminator.

The gene has its native promoter, introns and

terminator.(7)

Genetic elements

Allows construction of new genetic

combinations, introducing variability for gene

expression (1)

Involves both desired and undesired genetic elements of crossed plants

Involves exclusively the gene(s) of interest and no undesired genetic

elements (1)

Linkage drag Avoided (4) Present (8) Avoided (8)

Time factorTime saving since it is a

fast and precise tool(4)

Time- consuming and requires several

generations of breeding and selection

(5)

Time saving but takes much longer time than

intragenesis since genes/ fragmented genes may not be

readily accessible (5+8)

TechniquesMolecular Cloning

techniques: Recombination (5)

Crossing, mutagenesis and somatic

hybridisation (5)

Molecular techniques(5)

Differences between Traditional-, Intra- & Cis-genesis

Intragenesis Traditional breeding Cisgenesis

Safety

Deep concern about safety and impact on

health and environment (3)

Safe (crops being consumed since ages)

No environmental risk and safe as traditional

bred plants (3)

Gene pool Not conserved (3) Preserved (3) Preserved (3)

Genetic make up Original make up of plant is not maintained

Maintain original genetic make-up of plant variety

(3)

Maintain original genetic make-up of plant variety

(3)

Vigour of recipient plant

New gene modify vigour of target plant due to

gene influx from its wild relatives (8)

Change vigour, generating a change in the natural vegetation

(8)

No change in vigour resulting in no harm to

non-target species/ environmental hazards and potential allergens

(8)

Similarities between Trans, Intra & Cisgenesis & Conventional Breeding

● Integration of the genes requires chromosome breaks and natural DNA repair mechanisms.

● The process of creating modified plants aims at reducing the use of pesticides and insecticides which contribute to environmental problems .

Conclusion

According to the EFSA Panel on GMOs, ● similar hazards can be associated with cisgenic and

conventionally bred plants● novel hazards can be associated with intragenic and

transgenic plants

These breeding techniques can have variable frequencies and severities of unintended effects.The effects differ from breeding techniques and can not be predictable.Risks to human and animal health will depend on the exposure frequency; how the plant is cultivated and consumed.

References1) Espinoza, C., Schlechter, R., Herreta, D., Torres E., Serrano, A., Medina, C. and Arce-Johnson, P. (2013). Cisgenesis and Intragenesis: New tools for improving crops. Biological Research, 46, pp 323-331.2) FiBL. (2001). An evaluation for organic plant breeding. Plant Breeding Techniques.http://www.seedalliance.org/uploads/pdf/FiBL-PlantBreeding.pdf[Date accessed: 25.01.15]3) Holme, I.B., Wendt, T. and Holm, P.B. (2013). Current developments of intragenic and cisgenic crops. ISB news report, Aarhus University, Faculty of Science and Technology - Department of Molecular Biology and Genetics Research Centre Flakkebjerg, DK-4200 Slagelse, Denmark4) Holme, I.B., Wendt, T. and Holm, P.B. (2013). Intragenesis and cisgenesis as alternatives to transgenic crop development. Plant Biotechnology Journal, 11, pp 395-407.5) Hou, H., Atlihan, N. and Lu, Z-X (2014). New biotechnology enhances the application of cisgenesis in plant breeding. Frontiers in plant science, 5(389), pp 1-5.

References6) Podevin, N., Devos, Y., Liu, Y., Kärenlampi, S.O., Bradshaw, J., Jones, H., Kleter, G., Kuiper, H., Pöting, A., Sweet, J.B., Davies, H.V., et al. (2012). Scientific opinion addressing the safety assessment of plants developed through cisgenesis and intragenesis. EFSA Journal, 10(2), pp 1-33.7) Schouten, H.J. and Jacobsen, E. (2008). Cisgenesis and intragenesis, sisters in innovative plant breeding. Trends in Plant Science, 13(6), pp 260-261.8) Telem, R.S., Wani, S.H., Singh, N.B., Nandini, R., Sadhukhan, R., Bhattacharya, S. and Mandal, N. (2013). Cisgenics – A sustainable approach for crop improvement. Current Genomics, 14, pp 468-476.9) Wiel, C.V.D., Schaart, J., Niks, R., and Visser, R. (2010). Traditional Plant Breeding methods. Wageningen. http://www.rexresearch.com/agplntbrdg/tradplntbrdmethd.pdf[Date accessed: 25.01.15]

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