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Maneesh Mishra, Ph.D.Maneesh Mishra, Ph.D.
Biotechnology LaboratoryBiotechnology Laboratory
Central Institute for Subtropical HorticultureCentral Institute for Subtropical Horticulture
LucknowLucknow-- 227 107227 107
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Increase in Crop Yield
Green Revolution
Hybrids (Selective breeding Among
Sexually Compatible Partners)Use of Chemical Fertilizers
GREEN REVOLUTION
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TRADITIONAL BREEDING
teosinte
cob
corn
Wild tomato
(Lycopersicon pimpinellifolium )
D= 1 cm
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For years, framers have been selecting for certainfeatures in crops producing plants that are easierto grow, tastier, and bigger without knowing the
exact mechanism of how this occurs.
Nowadays scientists can transfer genesfrom one organism to another unrelatedorganism, producing what is now knownas genetically modified organism or transgenic animal/plantµ.
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DNA in chromosomes carries biological
information
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�� GenesGenes = the coding system for instructions
�� A g ene A g ene = is a segment of DNA
Guanine
(G)
Cytosine
(C)
Adenine
(A)
Thymine
(T)
bases DNA
DNA and GenesDNA and Genes
gene
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Genes are made of DNA
Promoter Protein coding region
Protein
Gene encode proteins. Proteins performs the biological functions
determining the characteristics of organism
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A gene is a DNA segment that encodes
a specific protein that contributes to
expression of a trait.
�The steps of the process:
mRNA molecule is transcribed accordingto the message encoded in the DNAcoding sequence (i.e., gene).
A protein is translated from the mRNAmolecule.
the production of this protein in the cellwill result in an expression of a trait.
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Genes encoding important traits
Extended shelf life
Insect, disease resistance
Uniform ripening,Increased sugars
Flavors
Antioxidants
Abiotic stress
Gene IdentificationGene Identification
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Gene manipulation
Genes are manipulated in many ways
Suppression ( a deleterious gene)
Over expression ( favorable gene)
Modification (enhancing new characteristics)
Transgene expression ( introducing new functions)
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How Genetic engineering technology works
A gene of known function (gene controlling fruit
ripening) is identified and isolated.
The gene must be stably incorporated in to plant.
The gene must be heritable.
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Marker gene Promoter TransgeneTermination
sequence
SelectionWhere
When
How much
What Stop
A DNA construct is used for transformationA DNA construct is used for transformation
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Genes are introduced in the plants using bacterial plasmid Genes are introduced in the plants using bacterial plasmid
as vectorsas vectors
Plasmids are circular molecules of DNA found in bacterial cell. They
confer survival of antibiotics
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How to get DNA in to plantsHow to get DNA in to plants
Agrobact erium
Particle bombardment
Electroporation
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Agrobact erium Agrobact erium tumef aci enstumef aci ens isis aa
naturallynaturally occurringoccurring bacteriumbacterium
thatthat hashas aa abilityability toto transfertransfer thethe
piecepiece of of DNADNA inin toto chromosomechromosome
TheThe abilityability hashas beenbeen usedused toto
transfertransfer DNADNA oror transformtransform plantsplants
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TTRANSFOR MATIONRANSFOR MATION
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Particle Bombardment System
R upture disc
Plant material
Stopping plate
R upture disc
Gold particle
Plant material
Stopping plate
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³Gene Gun´ Technique³Gene Gun´ Technique
DNA coatedgolden particles
Gene gun
Cell division
A plant cell with
the new gene
Transgenic plant
Plant cell
Cell¶s DNA
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CaMV Promoter OCS terminator
Bam H 1 Xba 1
CaMV PromoterNOS terminator
Bam H 1 Xba 1
pBinAR -12.2 Kb in sense orientatio
pBI121(13.5 Kb) in antisense orientation
GENE CONSTR UCT FOR PR SV
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83
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First transgenic plant
First Bt corn plants
First field tests
Delay-ripening tomato Commercialized in the US
Rotting resistant tomato approved by FDA
Herbicide resistant, insect resistant plants commercialized
GM maize approved by EU
HISTORY
OF TRANSGENIC
PLANTS
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Transgenic plum
Nontransgenic plum
Plum Pox Virus
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Flavr Saver Tomato: Extended shelf lifeFlavr Saver Tomato: Extended shelf life
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Transgenic tomatoes with
regulated ethylene production.
Delayed harvesting.
Fruits stays longer in the field.
Non-transgenic Transgenic tomato
Tomato with 4 with 2 colours
colours
g
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Tomato with reduced polygalucturoseTomato with reduced polygalucturose
Increased TSS in pasteIncreased TSS in paste
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Tomato plants artificially
inoculated with bacterial
speck disease
Plants on left has been genetically engineered
with gene for resistance to disease and plant onright has not.
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Transgenic tomato growing against salt stress
Transgenic grown
at x 40 salt
Non
transgenicgrown at x 40
salt
Non-
transgenic
grown at
200 mM
NaCl
Non-
transgenicgrown at 5
mMNaCl
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Delayed softening/ripening in transgenic apple
Non-Transgenic Transgenic
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Transgenic crops will create monsterTransgenic crops will create monster
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Transgenic cropsTransgenic crops--a curea cure--allall
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Many things are not natural. Good things result fromMany things are not natural. Good things result from
experimentation and improvement of nature:experimentation and improvement of nature:AntibioticsAntibiotics
AnaestheticsAnaesthetics
Warm housesWarm houses
And now, more nutritious foods!And now, more nutritious foods!
The testing of GM
foods is exemplary in its detail and takes atThe testing of GM
foods is exemplary in its detail and takes atleast four years of testing for it to be introduced to consumers.least four years of testing for it to be introduced to consumers.
GMOs have actually reduced the use of herbicides andGMOs have actually reduced the use of herbicides andpesticides.pesticides.
Now, is the time to experiment and not when a natural disaster Now, is the time to experiment and not when a natural disaster (Ex. overpopulation) plagues us and it is too late.(Ex. overpopulation) plagues us and it is too late.
TR UTH IS
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GENES IN THE SALAD
In the form of salad on an average we
eat: 100,000,000,000,000 (100 billion
genes)
(500 g Mixed Salad, Plant Cell: 50µm, Density:1g/cm,
~4x109 cells, 25,000 genes per cell)
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ThroughoutThroughout history,history, newnew scientificscientificdiscoveriesdiscoveries havehave challengedchallenged
conventionalconventional thinkingthinking (Galileo(Galileo--
CopernicusCopernicus theory,theory, SunSun--EarthEarth
Movement)Movement) However,However, over over thethe yearsyearssocietysociety hashas learnedlearned toto acceptaccept indeed,indeed,
celebratecelebrate thesethese discoveriesdiscoveries andand socialsocial
andand intellectualintellectual benefitsbenefits theythey havehaveconveyedconveyed..
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Transgenic cropsTransgenic crops--A bountifulA bountiful
harvestharvest
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± Patents, plant breeder¶s rights, tr ade secrets, tr ade
marks, copy rights
± Gene constructs would be covered by patents
IPR in Biotechnology
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A gene construct is built from
genetic material isolated from
different organisms
± E.g. promoter from CaMV, a bacterial DNA vector
( Agrobacterium plasmid), one or more genes th
atmay have been modified in the labor atory,
termination and signalling sequences and a
selectable marker gene.
� Involves both Process and Product
Gene Construct
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An exclusive right gr anted for an invention,
which is a product or a process that
provides, in gener al, a new way of doing
something, or offers a new technical
solution to a problem.
----------------------------------------------------------------
Once issued, a patent gives the inventor the legal right to create a
limited monopoly by excluding others from creating, producing,
selling or importing the invention.
This right is of limited dur ation, for a minimum period of 20 years
Patent
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Usefulness or industrial application
Newness or novelty
Non-obviousness or inventive step
Conditions for granting Patents
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Be careful while obtaining permission to
use the constructs for research
N ote
The high incidence of the use of the promoter CaMV /35S
The use of genes with very specific characteristics
like the Cry genes
The use of selectable markers such as GUS,
kanamycin and hygromycin resistance genes. The basic constructs containing selectable markers,
promoters, and genetic markers
Patent vis a vis Gene
Constructs
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Be aware
Of the principles of territoriality of patents, and, as a consequence, of the
implications for the application of
their research.
Of the fact that whether a gene
construct is patented or not and if
patented, about the need to seek thelicense.
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Pl ease note
Gene constructs may not be patentedin our country, and therefore the
original patent may not be important
for the academic research purpose.
However, the actual implications
depend on the final use of the
research results e.g. a transformed
plant.
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Mostly the gene constructs are notobtained by a formal contractual
arrangement ("lacking written
approval", "international collaboration",
and "not known").
Furthermore, the use of licenses is low,
in contrast to the use of non-writtenagreements and international
collaboration.