Production of transgenics in oilseeds by Kanak Saxena
1. Department of Plant Breeding and
Genetics
CREDIT SEMINAR
ON
Jawaharlal Nehru Krishi Vishwa Vidyalaya
Jabalpur (M.P.)
(2015-16)
“PRODUCTION OF TRANSGENICS IN
OILSEED CROPS”
Guided by Speaker
Dr.(Mrs.) Rajani
Bisen
Kanak Saxena
Roll no. 234
1
2. Introduction
Why transgenic production
Need of transgenics in oilseeds
Steps of transgenic Production
Methods of Transformation
Transgenic Oilseed crops (Case studies)
Application
Limitations
Conclusion
CONTENT
2
3. • India is the fourth largest oilseed producing country in the world
• India has the largest area and production of oilseeds in the world
• Oilseeds are rich sources of energy and nutrition.
• Oilseeds also contain carbohydrates, vitamins and minerals
• Oilseeds and oilseed meals have an important role in relieving the
malnutrition and calorie nutrition of human and animal population. In
addition, the vegetable oils are useful as lubricants, surface coatings,
cosmetics and as raw material for various industrial products like,
paints, varnishes, hydrogenated oil, soaps, perfumery, lubricants, etc
• Oil-cake which is the residue after the oil is extracted from the
oilseeds, forms an important cattle-feed and manure
INTRODUCTION
3
4. • The most important annual oilseed
crops are
1. Groundnut
2. Rapeseed-Mustard
3. Sesame
4. Sunflower
5. Safflower
6. Soybean
7. Niger
8.Castor and
9. Linseed.
4
5. Area, Production and Productivity
Source --Department of Agriculture and cooperation DAC (2013-14)
AREA
(lakh ha)
285.25
PRODUCTION
(million tonne)
328.77
PRODUCTIVITY
(Kg/ha)
1094
5
8. Why transgenic production ?
Rapid method of crop improvement
Overcome problems related to wide hybridization
Evolution of New Genotypes
Effectiveness
Generates useful genetic variation
8
9. Survey says-
• Reasons for growing transgenics:
–Weed control
–Better yield, more return, profit
–Reduce costs
–Less time
9
10. Survey says-
• Reasons for not growing transgenics:
–Overall costs
–No need to change
–Market access
10
12. Need of transgenics in oilseeds?
• India is a leading importer of vegetable oil so we
have to develop some improved varieties so we can
able to break all constraints in oilseed production
• Although we have wide variability in oilseeds ; wild
as well as cultivated but we are not able to mitigate
the requirement of vegetable oil in our country
• So there is a need to create variability in the existing
population in the existing crop and for this creation
of variability transformation method is the only
milestone in the improvement of oil production
12
13. Sr.
No. Crop Trait Countries where approved
1 Sunflower Herbicide tolerance Canada
2
Argentine
canola
Herbicide tolerance and
improved protection against
weeds
Canada, USA,
3 Soybean
Improved weed control and
herbicide tolerance, increased
cooking quality
USA, Argentina, Japan,
Canada, Uruguay, Mexico,
Brazil and South Africa, Czec
Republic, European Union,
Korea, Russia, Switzerland,
Taiwan, U.K., Philippines
and Australia
4
Flax,
Linseed
Herbicide tolerance, antibiotic
resistance and improved weed
protection
Canada, USA
Worldwide Transgenic crops approved for commercial use
Source ; the Indian Society of Oilseeds Research,DOR, Hyderabad (2013)
13
14. Institute Plants/
crops
Transgene(s)
inserted
Aim of the project
Indian Agricultural
Research, Institute, New
Delhi
Mustard/
rapeseed
Arabidopsis
annexin gene
To generate stress-tolerant
plants
Indian Agricultural
Research, Institute, New
Delhi
Mustard/
rapeseed
Choline
dehydrogenase
To generate abiotic stress-
tolerant plants
Delhi University, South
Campus, New Delhi
Mustard/
rapeseed
Bar, Barnase,
baraster
To generate herbicide-tolerant
plants, male sterile and
restorer lines for hybrid seed
production
Indian Agricultural
Research, Institute, New
Delhi
Brassica Chitinase,
glucanase and
RIP
To generate plants
resistant to fungal attack
Major Developments in Transgenic Research and its Applications
in Public Sector
Source ; the Indian Society of Oilseeds Research,DOR, Hyderabad (2013)
14
15. Institute Plants/
crops
Transgene(s)
inserted
Aim of the project
Proagro PGS
(India) Ltd,
Gurgaon
Brassica
/
mustard
Bar, barnase,
barstar
To develop
superior hybrid
cultivars
Tata Energy
Research
Institute, New
Delhi
Mustard Ssu-maize
Psy and Ssu-
tpCrt1 gene
To generate plant
containing high
level of b-carotene
Major Developments in Transgenic Research and its Applications
in Private Sector
Source ; the Indian Society of Oilseeds Research,DOR, Hyderabad (2013)
15
16. Steps for developing transgenic crops
o Identification of gene
o Gene transfer
o Regeneration from callus/tissue/protoplast
o Gene expression to the desired level
o Backcross to produce varieties
o Field test
o Approval for commercialization
16
18. Vector Based Method:- Infecting plant cells
with plasmid as vector carrying the desired gene
1) Agrobacterium tumefaciens
2) Agrobacterium rhizogenes
Direct Gene Transfer Method :-
Shooting microscopic particle containing gene
directly into the Cell
1. Particle Bombardment (Gene Gun)
2. Electroporation of protoplast
3. Microinjection
4. Liposome mediated gene transfer
Direct & Indirect Methods of Gene Transfer
18
21. To identify cells/tissues in which new genes are
incorporated into plant’s DNA, grow in media containing
antibiotics or herbicides.
Successful
transformant
Identification of Transformed cell
21
22. Whole plants with
inserted genes are
regenerated through
tissue culture.
Regeneration of Transformed cells
22
23. 1. Electroporation –
Electrical impulses are used to increase membrane and cell wall
permeability to DNA contained in the surrounding solution.
Direct MethodsDirect Methods
23
24. 2. Microinjection - injection of DNA directly into the cell
Nucleus using an ultrafine needle.
2. Microinjection - injection of DNA directly into the cell
Nucleus using an ultrafine needle.
24
26. 4.Polyethelyne glycol –
Plant cell protoplasts treated with PEG are
momentarily permeable, allowing uptake of
DNA from the surrounding solution.
26
27. • Resistant to herbicide – eg. Soybean, Linseed
• Resistant to virus – eg. Sunflower
• Production of male sterile lines – eg. Brassica spc.
• Improved oil quality and quantity – eg. Canola
• Improved nutritional quality – eg. Canola
Applications Of Transgenic In Oilseeds
27
30. A mutant aro gene from bacteria Salmonella typhimurium has
been used for developing tolerance to glyphosate.
The target site of glyphosate is a chloroplast enzyme 5-enol
pyruvylshikimic acid 3-phosphate synthase (EPSPS)
Introduction to mutant aro gene produces modified EPSPS, not
recongnizable to glyphosate.
Development of herbicide resistant crops allows the elimination
of surrounding weeds without harm to the crops.
Roundup Ready Soybeans; The first variety was also known as
GTS 40-3-2 are a series of genetically engineered varieties of
glyphosate resistant soybeans produced by Monsanto.
Steinrucken, H.C et al., 1980
30
32. Genetic transformation of cultivated sesame through particle
bombardment using 5-day-old apical,meristematic tissues of
germinating seedlings
• An in vitro plant generation and genetic transformation
protocol was established in sesame (sesamum indicum l. cv
rama) through biolistic particle gun bombardment.
• 5-day-old apical, meristematic tissues of in vitro-
germinating seedlings were used as explants.
• A synthetically designed bialaphos resistance gene
(bar)was used for transformation.
Bhattacharyya, et al., 2015
32
35. Generation of white mold disease-resistant sunflower
plants expressing human lysozyme gene
Generation of white mold disease-resistant sunflower
plants expressing human lysozyme gene
• Gene used - human lysozyme gene
• Method used - Agrobacterium mediated transformation
• The plasmid pNGL was used for transformation of Agrobaterium
tumefaciens. It contains the structural gene for neomycin
phosphotransferase-II npt II, which encodes resistance to the
antibiotic kanamycin sulphate, the β-glucuronidase gus reporter gene
and the human lysozyme gene.
• The expression of the human lysozyme gene is under the control of
the cauliflower mosaic virus (CaMV) 35S promoter and is terminated
by the nopaline synthase Nos gene terminator.
• Southern blot analysis, Western blotting ,Northern blot analysis
Sawahel, et al.,(2006)
35
37. Jagannath, et al.,2012
The barnase/barstar gene system in the GM mustard
• Barnase/barstar gene system, in which herbicide resistance is linked
with male sterility, so that the herbicide will kill the male fertile lines,
leaving the seed producing male sterile plants unharmed.
• Barnase, a ribonuclease (an enzyme) from bacillus amyloliquefaciens,
inhibits pollen formation and results in male sterility in the transgenic
plants.
• The bar gene from streptomyces hygroscopicus encodes for the
enzyme phosphinothricin acetyltransferase, that restores male fertility
in the transgenic lines.
• A phsphonithricin resistance-coding (pat) gene is used to eliminate
undesirable segregates, by spraying an herbicide. 37
40. Improved rapeseed
• Improved rapeseed cultivars were free of erucic acid
and glucosinolates. Erucic acid tastes bitter and had
prevented the use of rapeseed oil in food.
• Gluconsinolates, which were found in rapeseed meal
leftover from pressing, are toxic and had prevented
the use of the meal in animal feed. These new
cultivars are known as "double-zero" rapeseed. In
Canada, where "double-zero" rapeseed was
developed, the crop was renamed "canola" (Canadian
oil) to differentiate it from non-edible rapeseed.
Bin jhu, et al.,(2011)
40
41. Laurate canola oil
• GM rapeseed enriched with lauric acid can be used for
producing fat-based coatings in food processing
• Canola plant modified with thioesterase gene obtained
from California bay laurel tree
• Enzyme produces lauric acid from genetically
modified (GM) canola seeds
• Low saturated fat content
• Heat tolerant
• Does not break down
• Excellent for high temperature cooking processes
• 41
43. Various varieties of oilseeds have been developed
through Transgenic for different traits and released for
commercial cultivation.
Genetic engineering offers considerable potential for
genetic improvement of crop plants, especially for
disease and pest resistance, and improved quality
characteristics.
The Transgenic technology has great potential to
generate new varieties along with the conventional
breeding.
CONCLUSION
43
44. References
• Ag-West Biotech, “Canola- Biotechnology’s Powerhouse
Crop,” AgBiotech Infosource Issue 21, May , 1996
http://www.agwest.sk.ca/event_inf_may96.shtml>, April,
2002.
• Department of Agriculture and cooperation DAC (2011-12)
• FAOSTAT 2012
• Jagannath, Arun ; Arumugam, N. ; Gupta, Vibha ; Pradhan,
Akshay ; Burma, Pradeep Kumar ; Pental,
Deepak (2002)Development of transgenic barstar lines and
identification of a male sterile (barnase)/restorer (barstar)
combination for heterosis breeding in Indian oilseed mustard
(Brassica juncea) Current Science, 82 (1). pp. 46-52
• Steinrücken, H.C.; Amrhein, N. (1980). "The herbicide
glyphosate is a potent inhibitor of 5-enolpyruvylshikimic acid-
3-phosphate synthase". Biochemical and Biophysical Research
Communications 94 (4): 1207–12.
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