1. Applications Of Genetic Engineering
Techniques In Agriculture
B.DEVADATHA
123680029
m.Sc biochemistry and

2. Why genetically engineer plants?
• To improve the agricultural, horticultural or
ornamental value of a crop plant
• To serve as a bioreactor for the production of
economically important proteins or metabolites
• To provide a powerful means for studying the
action of genes (and gene products) during
development and other biological processes

3. Genetic engineering techniques applied to plants
METHOD SALIENT FEATURES
1.VECTOR MEDIATED GENE
TRANSFER
a. Agrobacterium mediated gene
transfer
b. Plant viral vectors
Very efficient but limited to a selected group of plants
Ineffective, hence not widely used
2.DIRECT OR VECTORLESS
DNA TRANSFER
a. Electroporation
b. Microprojectile
c. Liposome fusion
d. Silicon carbide fibres
Mostly confined to protoplasts that can be regenerated to viable
plants
Limited use only one cell can be microinjected at a time
Confined to protoplasts that can be regenerated into viable whole
plants
Requires regenerable cell suspensions
3 CHEMICAL METHODS
a. Polyethylene glycol mediated
b.Diethylaminoethyl(DEAE)dextra
n- mediated
Confined to protoplasts. Regeneration of fertile plants is frequently
problematical
Does not result in stable transformants

4. AGROBACTERIUM MEDIATED GENE TRANSFER
• Agrobacterium tumefaciens is a naturally occurring
Gram negative bacteria
• Causes Crown gall disease by transferring T-DNA from
its Ti plasmid in variety of dicotyledonous plants
• Plants infected with this bacterium develop tumour like
swellings galls that typically occur at crown of plant just
above soil level
• Following infection ,the bacterium transfers part of its
DNA into plant & this DNA integrates into plant genome
causing production of tumors and associated changes
in plant metabolism

5. Process of Infection
• A.tumefaciens infects plants through wounds
• The motile bacterial cells are attracted to
wound site by phenolic compounds such as
acetosyringone secreted at wound site
through specific chemotactic receptors
present in bacterial membrane
• Acetosyringone at higher concentrations
activates Vir genes leading to production of
proteins that are inserted into bacterial cell
membrane for upake of opines produced by
tumors
• Acetosyringone also causes production of an
Endonuclease that encodes a part of Ti
plasmid termed T-DNA

6. Gene gun/Biolistic bombardment system
 Desired DNA is coated on microscopic gold or tungsten beads
 The particles are fired by a gun into plant tissue and penetrate the cell wall
 DNA unwinds from gold carrier
particle and enters the nucleus

7. Applications of Plant Genetic Engineering
• Improved Nutritional Quality
• Insect resistance
• Disease resistance
• Herbicide resistance
• Salt tolerance
• Delayed Fruit Ripening
• Biopharmaceuticals and Vaccines

8. Improved Nutritional Quality
• 124 million children worldwide are deficient in vitamin A, which leads to death
and blindness
• b-carotene is precursor to vitamin A & consuming milled rice leads to vitamin A
deficiency
• Mammals make vitamin A from b-carotene,
a common carotenoid pigment normally found
in plant photosynthetic membranes
Daffodil phytoene synthase gene
carotene desaturase gene
Daffodil lycopene b-cyclase gene
Endogenous human gene

9. RAINBOW CAULIFLOWER
• Produced by traditional breeding –Non
Transgenic
• The Orange cauliflower has higher
than normal levels of b-carotene
that encourages healthy skin
• Purple colour comes from
Anthocyanin which may prevent
Heart disease by slowing blood clotting
• Tests of the orange cauliflowers in
America found that they contained 25
times the concentrations of beta
carotene in normal cauliflowers

10. Insect resistance
• It is estimated about 15% of world’s crop yield is lost
through insects
• Bacillus thuringiensis is a Gram negative ,soil bacterium
produces parasporal crystalline proteinous toxin with
insecticidal activity
• When the Bt toxin gene was introduced into economically
imp crop plants they develop resistanace for major insects
obviating the need for use of insecticides
• Bt toxin opens cation selective pores in membranes
,leadng to inflow of the cations into cells that causes
osmotic lysis & destruction of epithelial cells
• Bt genes could be expressed
• in all parts of plants

11. Disease resistance plants
• Genes that provide resistance against plant
viruses have been successfully introduced
into crop plants such as tobacco, tomato,
rice, potato ,etc
• Transgenic tobacco plant
• Expressing tobacco mosaic virus coat protein
gene were first developed
• Viral capsids inhibit viral replication of TMV when infected
• Virus coat protein mediated protection is
successful for viruses with ss RNA

12. Herbicide resistance
• Genes for resistance against certain Herbicides have
been introduced into crop plants so they can thrive even
when exposed to Herbicides
• Glyphosate is a non selective Herbicide that inhibits 5-
enolpyruvylshikimate-3-phosphate synthase (EPSP) a
chloroplast enzyme in the shikimate pathway and plays a
key role in the synthesis of aromatic amino acids such as
tyrosine and phenylalanine
• Herbicides Sulfonylureas & Imidazolines inhibits
Acetolacetate (ALS),key enzyme in synthesis of Isoleucine
,Leucine & Valine
• Some other Herbicide resistant plants also developed
eg:bromoxynil,atrazine ,phenocarboxylic acids

13. Salt tolerance
• A large fraction of world’s
irrigated land cannot be
used to grow most
important crops due to
increased salinity in soil
• Resarcher’s have created
transgenic tomatoes that
grew well in saline soils
• The transgene introduced
was sodium/proton
antiport pump that
sequestered excess
sodium in vacuole of leaf
cells

14. Flavr savr tomato
• Most tomatoes that have to be shipped to
market are harvested before they are ripe
otherwise, Ethylene synthesized by tomato causes
them to ripen & spoil before they reach customer
• Flavr savr tomatoes have been constructed that
Express an antisense mRNA complementary to
mRNA for an enzyme involved in ethylene
Production
These tomatoes make only 10% of normal amount
of enzyme thus delaying ethylene production

15. Biopharmaceuticals and Vaccines
• Expression of Hepatitis B Surface
Antigen in Transgenic plants
• Tobacco plants were genetically
transformed with gene encoding
Hepatitis B surface antigen
(HBsAg) linked to a nominally
constitutive promoter were
generated
• Recombinant HBsAg purified
from transgenic plant had
properties similar to Human
serum derived HBsAg

16. Edible vaccines
• Vaccines consisting of transgenic plant-derived antigens offer
a new strategy for development of safe, inexpensive vaccines.
• The vaccine antigens can be eaten with the edible part of the
plant or purified from plant material
Rabies- Tomato plants expressing rabies antigens could induce
antibodies in mice
Cholera-Transgenic potato with CT-B gene of Vibrio
cholerae was shown to be efficacious in mice .
Norwalk virus- transgenic potato expressing norwalk virus
antigen showed seroconversion
Hepatitis B- First human trials of a potato-based vaccine
against hepatitis B have reported encouraging results
If vaccines are intimately presented together with food, the
guts immune system faces a conundrum

17. What are some of the advantages of GM
foods?
GM crops are more productive and have a larger yield.
Offer more nutritional value and better flavor.
A possibility that they could eliminate allergy-causing properties in some
foods.
Inbuilt resistance to pests, weeds and disease.
More capable of thriving in regions with poor soil or adverse climates.
More environment friendly as they require less herbicides and
pesticides.
Foods are more resistant and stay ripe for longer so they can be shipped
long distances or kept on shop shelves for longer periods.
As more GM crops can be grown on relatively small parcels of land, GM
crops are an answer to feeding growing world populations.

18. What are some of the advantages of GM foods?
• Disease resistance There are many viruses, fungi
and bacteria that cause plant diseases.
• Plant biologists are working to create plants
with genetically-engineered resistance to these
diseases.
• Cold tolerance Unexpected frost can destroy
sensitive seedlings. An antifreeze gene from cold
water fish has been introduced into plants such
as tobacco and potato.
• With this antifreeze gene, these plants are able
to tolerate cold temperatures that normally grow

19. Environmental & health impacts of GM
crops
• Toxic to harmless non-target species. Long-term
exposure to pollen from GM insect resistant maize
causes adverse effects on the behaviorand survival
of the monarch butterfly
• Toxic to beneficial insects. GM Bt crops adversely
affectbeneficial insects important to controlling
maize pests, such as green lacewings
• A threat to soil ecosystems. Many Bt crops
secrete their toxin from their roots into the soil.
Residues left in the field contain the active Bt toxin
• Risk for aquatic life. Leaves or grain from Bt maize
can enter water courses where the toxin can
accumulate in organismsand possibly exert a toxic
effect

20. Environmental & health impacts of GM
crops
• Herbicide-tolerant crops may cross-pollinate
weeds, resulting in "superweeds"
• Certain gene products may be allergens, thus
causing harm to human health
• recent study found the Cry1Ab Bt toxin in the
blood of pregnant women and their fetuses
showing that it can cross the placental
boundary
• Studies indicate Roundup may be toxic to
mammals and could interfere with hormones
• There may be unintended harm to wildlife and
beneficial insects

21. conclusion
• Genetically-modified foods have the potential to solve many of the
world's hunger and malnutrition problems, and to help protect and preserve the
environment by increasing yield and reducing reliance upon chemical pesticides
and herbicides.
• Yet there are many challenges ahead for governments, especially in the areas of
safety testing, regulation, international policy and food labeling.
• Many people feel that genetic engineering is the inevitable wave of the future
and that we cannot afford to ignore a technology that has such enormous
potential benefits.
• However, we must proceed with caution to avoid causing unintended harm to
human health and the environment as a result of our enthusiasm for this
powerful technology.
• At the present time, genetically modified foods are dangerous for the world
that we live in, not just for its people, but the delicate balance between
organisms that inhabit it.

22. Sources
• GM Science Review First Report (http:/ / www. bis. gov. uk/ files/
file15655. pdf), Prepared by the UK GM Science Review panel
(July2003).
• Maxmen, Amy (2 May 2012) First plant-made drug on the market (http:/
/ blogs. nature. com/ news/ 2012/ 05/first-plant-made-drug-on-the-
market. html) Nature, Biology & Biotechnology, Industry. Retrieved 26
June 2012
• Conner AJ, Glare TR, Nap JP. The release of genetically modified crops
into the environment. Part II. Overview of ecological risk assessment
(http:/ / www. ncbi. nlm. nih. gov/ pubmed/ 12943539) Plant J. 2003
Jan;33(1)
• Vaccine antigen production in transgenic plants: strategies,gene
constructs and perspectives Elsevier.com
• Principles of Gene Manipulation and Genomics Sandy B.
Primrose, Richard Twyman