application of tissue culture in crop improvement (transgenic plants)

1. APPLICATION OF TISSUE CULTURE IN CROP
IMPROVEMENT
Presented by,
M.Uma Maheshwari

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. TRANSGENIC PLANTS
NUTRITIONAL
QUALITY
BIOTIC STRESS
TOLERANCE
ABIOTIC STRESS
TOLERANCE
PHARMACEUTICALS
& EDIBLE VACCINE
HYBRID DEVELOPMENT
FOR HIGHER YIELD
ENHANCED
SHELF LIFE
INDUSTRIAL
PRODUCTS

4. Applications of Tissue Culture
• Improved Nutritional Quality
• Insect resistance
• Disease resistance
• Herbicide resistance
• Salt tolerance
• Delayed Fruit Ripening
• Biopharmaceuticals and Vaccines

5. Golden Rice
• 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

6. The prototype of golden rice was developed in 2000 and is a
light yellow color (b). It contains 1.6 mg/g of carotenoid.
In 2005, new transgenic lines were developed that dramatically
increased the amount of carotenoid synthesized, making the
rice a deep golden color (c).
This latest form contains 37 mg/g of carotenoid, of which
84% is b-carotene – trial

8. Rainbow cauliflower
• Produced by traditional breeding –Non Transgenic
• The Orange cauliflower has higher than normal
levels of b-carotene that encourageshealthyskin
• Purple colour comes from Anthocyanin which may
prevent Heart disease by slowing blood clotting
• Tests of the orange cauliflowersin America found
that they contained25 times the concentrationsof
beta carotenein normal cauliflowers

9. • Bt gene of a bacterium Bacillus
thuringiensis has found to encode endotoxin
which pose cidal effect on certain insect
pests
• The cry gene found to express the
proteinaceous toxin, when specific pest
ingest the toxin, they are killed
• The toxin affect specific group of insects
and do not harm silkworm, butterflies and
other beneficial insects
• Using biotechnological approaches many
transgenic plants with cry gene have been
developed eg., Bt-brinjal, cauliflower,
cabbage, canola, corn, maize, tobacco, rice,
soyabean

10. δ -endotoxin gene (Cry gene) of Bacillus thuriengenesis
GENE FOR Bt TOXIN WAS TRANSFERRED
TO OBTAIN BT TRANSGENIC PLANTS
PLANT SYNTHESIZES INACTIVE PROTOXIN
PROTEINASE
DIGESTION IN
INSECT GUT
MAKES THE
ACTIVE TOXIN
Toxin binds a receptor on the gut epithelial cells, forms a channel
on the membrane. This causes electrolyte leakage and insect death
INSECT FEEDS ON
TRANSGENIC PLANT

11. • Plantsthat can tolerateherbicides
• The herbicide disturb the metabolic
activity of photosynthesisor synthesis
of amino acid
• For the development of herbicide
resistantplants two main strategiesare
being applied
– Modification of target molecules that may
be insensitive to herbicides
– Degradation of herbicides

12. • Attempts have been made to develop resistant against three herbicides
– Glyphosate
– Sulphonylurea
– Imidazolinoles
• A herbicide resistant gene for EPSPS was isolated from plants and transferred
to Petunia and transgenic Petunia was developed
• Transgenic tomato was developed by introducing a mutant als gene of
tobacco (inhibits sulphonylurea, imidazolinoles)
• A gene resistant to PPT was isolated from Medicago sativa inhibits GS
involved in ammonia assimilation
• Incorporated in to tobacco and thus transgenic tobacco is developed with PPT
resistant

13. • A number of micro organismsare also
involved in the degradationof herbicides
• Accomplished by genes coding specific
enzyme PAT which degrades the
herbicide PPT
• Nitrilaseencoded by bxn gene of
Klebsiellabromoxynil, GST degrades
herbiscide Atrazine
• Several crops have developed:
transgenic potato, oil seed rape and
sugarbeet(withbar gene), transgenic
tomato (bxn gene)

14. • Plant viruses yield several loss in economically important plants
• Two main approaches for developing genetically resistant plants
– PDR (PathogenDerived Resistance)
– non PDR (non- pathogenderived resistance)
• Roger Beachy and co-workers first introduced Coat protein (CP) of TMV in to
tobacco
• In many crops, virus resistance have been achieved through introducing CP
• CPMR is the most favoured strategy to make virus resistant plants

15. Host Transgene Increase in yield
over non-transgenic
plants
Tomato TMV, CP gene
CMV, Satellitegene
40
14
Potato PVX, PVY CP 38
Squash CMV+ZYMV+WMV2 +CP
ZYMV+WMV2 CP
97
90
Papaya PRSV CP 90

16. Disease resistance plants
• Genes that provide resistanceagainstplant
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
successfulfor viruses with ss RNA

17. • A large fraction of world’s irrigated
land cannot be used to grow most
important crops due to increased
salinity in soil
• Researcher'shave 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

18. Delayed ripening
• Antisense technology is used
produce the Flavr-Savr tomato in
1994.
• Enzyme polygalacturonase
breaks down structural
polysaccharide pectin in wall of a
plant.
• This is part of the natural decay
process in a plant
• Monsanto identified the gene than
encodes the enzyme and made
another gene that blocked the
production of the enzyme.

19. • Genetically modified plants have been used as
“bioreactors” to produce therapeutic proteins A
recent contribution is the generation of edible
vaccines.
• Edible vaccines are vaccines produced in
plants that can be administered directly
through the ingestion of plant materials
containing the vaccine. Eating the plant would
then confer immunity against diseases.
• Edible vaccines produced by transgenic plants
are attractive for many reasons.
• The first human clinical trial took place in
1997. Vaccine against the toxin from the
bacteria E.coli was produced in potato

20. Edible vaccines
• Vaccines consisting of transgenicplant-derivedantigens
offer a new strategyfor development of safe,
inexpensive vaccines.
• The vaccine antigens can be eaten with the edible part of
the plant or purifiedfrom 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 efficaciousin mice .
 Norwalkvirus- transgenicpotato expressing norwalk
virus antigen showed seroconversion
 HepatitisB- First human trialsof a potato-based vaccine
against hepatitisB have reported encouraging results
 If vaccines are intimatelypresented together with food,
the guts immunesystem faces a conundrum

21. • Plant seeds may be a potential source for
plasticsthat could be producedand easily
extracted.
• A type of PHA (polyhydroxylalkanoate)
polymer called “poly-beta-
hydroxybutyrate”, or PHB, is producedin
Arabidopsis, or mustard plant.
• PHB can be made in canola seeds by the
transferof three genes from the bacterium
Alcaligenes eutrophus, which codes for
enzymes in the PHB synthesispathway.
• A polymer called PHBV producedthrough
Alicaligenesfermentation, which is sold
under the name Biopol

22. 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.

23. What are some of the advantages of GM foods?
• Disease resistance Thereare many viruses,
fungi and bacteriathat cause plant diseases.
• Plant biologistsare working to create
plants with genetically-engineered
resistanceto these diseases.
• Cold tolerance Unexpected frost can
destroy sensitiveseedlings. An antifreeze
gene from cold water fish has been
introduced intoplants such as tobacco and
potato.
• With this antifreezegene, these plantsare
able to tolerate cold temperatures that
normally grow

24. • 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.