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Genetic transformations- herbicide resistant plants
Ashwin Jayale Id no-PALB 1222
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Contents Introduction Herbicide resistant plants. Herbicide and mode of action on plants. Genetic engineering approach. GM plants and statistical data Case studies Controversies Conclusion
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Introduction In molecular biology transformation is the genetic alteration of a cell resulting from the
direct uptake, incorporation and expression of exogenous genetic material (exogenous
DNA) from its surroundings and taken up through the cell membrane(s).
Genetically modified foods or biotech foods are foods derived from genetically
modified organisms (GMOs).
Genetically Modified foods (GM foods) were first put on the market in 1996. Typically,
genetically modified foods are transgenic plant products: soybean, corn, canola, rice, and
cotton seed oil.
The first commercially grown genetically modified whole food crop was a tomato
called FlavrSavr
Afterwards in 1995, a biotech company Monsanto introduced herbicide immune soybean
also known as the Roundup ready .
In 1996, the first genetically modified Canola was available on the market.3
Herbicide resistant plants Herbicide resistance: "Herbicide resistance is the inherited
ability of a plant to survive and reproduce following exposure
to a dose of herbicide normally lethal to the wild type.
BY WASSA.
In a plant, resistance may be naturally occurring or induced
by such techniques as genetic engineering or selection of
variants produced by tissue culture. Source: Weed
Technology Volume 12, Issue 4 (October-December) 1998. p.
789. weed science society of America.
Taking in the entire crop-growing period from 1996 to 2004,
the US used some 62.7 million kg more herbicides on
herbicide-resistant crops compared with conventional
management (Benbrook 2004).
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Importance Excessive weed growth forces crops to compete for sunlight and nutrients, often leading to
significant losses. Because herbicides cannot differentiate between plants that are crops and
plants that are weeds, conventional agricultural systems can only use 'selective' herbicides.
Such herbicides do not harm the crop, but are not effective at removing all types of weeds
If farmers use herbicide resistant crops, 'non-selective' herbicides can be used to remove all
weeds in a single, quick application.
Herbicide resistant crops also facilitate low or no tillage cultural practices, which many
consider to be more sustainable.
'Broad-spectrum', or non-selective herbicides are effective at killing a wide range of
weeds. The problem is, they can also kill valuable crops.
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Food Properties of the genetically modified variety Modification
Soybeans Resistant to glyphosate herbicides
Herbicide resistant gene taken from bacteria inserted into
soybean Corn, field (Maize) Resistant to glyphosate
herbicides. New genes, some from the bacterium Bacillus thuringiensis,
Alfalfa Resistant to glyphosate herbicides
New genes added/transferred into plant genome.
Hawaiian papaya Variety is resistant to the papaya ringspot virus.
New gene added/transferred into plant genome
Tomatoes
Variety in which the production of the
enzyme polygalacturonase (PG) is suppressed, retarding fruit softening after harvesting.
A reverse copy (an antisense gene) of the gene
responsible for the production of PG enzyme added into plant
genome
CanolaResistance to herbicides
(glyphosate or glufosinate), high laurate canola
New genes added/transferred into plant genome
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Herbicide and mode of action Glyphosate (N-phosphonomethyl-glycine) is the most
widely used herbicide in the world: glyphosate-based
formulations exhibit broad-spectrum herbicidal activity .
The extraordinary success of this simple, small molecule
is mainly attributable to the high specificity of glyphosate
for the plant enzyme enolpyruvyl shikimate-3-phosphate
synthase in the shikimate pathway, leading to the
biosynthesis of aromatic amino acids.
herbicide glyphosate (N-phosphonomethyl-glycine) has
had the greatest positive impact. Developed by the
Monsanto Co. and introduced to world agriculture in
1974.7
Shikimmic acid pathwayGlyphosate works by blocking the plants' ability to produce certain proteins and it disrupts amino acid synthesis. It blocks the shikimic acid pathway found only in certain plants .
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Roundup is the brand name of a systemic, broad-spectrum herbicide produced by the U.S. company Monsanto, and contains the active ingredient glyphosate.
Monsanto developed and patented the glyphosate molecule in the 1970s, and marketed Roundup from 1973.
Herbicide properties
Main active ingredient=isopropylamine salt of Glyphosate
Mode of action=5-enolpyruvylshikimate-3-phosphate synthase(EPSPS) inhibitor
Monsanto also produces seeds which grow into plants genetically engineered to be tolerant to glyphosate, which are known as Roundup Ready crops.
The genes contained in these seeds are patented.
Soy was the first Roundup Ready crop, and was produced at Monsanto's Agracetus Campus located in Middleton, Wisconsin.
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Genetic engineering approach Genetic engineering, also called genetic modification,
is the direct human manipulation of an
organism's genome using modern DNA technology.
An organism that is generated through the introduction
of recombinant DNA is considered to be a genetically
modified organism.
It involves the introduction of foreign DNA or synthetic
genes into the organism of interest.
GMOs Express traits not normally found in nature
Result of introducing foreign DNA
Highly controversial
Safety concerns
Environmental implications.10
Mechanism GM herbicide resistant crop Some microorganisms contains genes for producing 5-enolpyruvylshikimate-3-phosphate
synthase (EPSPS);5-enolpyruvylshikimate-3-phosphate synthatase ; phosphoenolpyruvate:3-
phosphoshikimate 5-O-(1-carboxyvinyl)-transferase these resistant to glyphosate inhibition.
These used in genetically modified crops, and isolated from Agrobacterium strain CP4 (CP4
EPSPS) that was resistant to glyphosate.
The CP4 EPSPS gene was cloned and inserted into soybeans. The CP4 EPSPS gene was
engineered for plant expression by fusing the 5' end of the gene to a chloroplast transit
peptide derived from the petunia EPSPS.
The plasmid used to move the gene into soybeans was PV-GMGTO4.
It contained three bacterial genes, two CP4 EPSPS genes, and a gene encoding beta-
glucuronidase (GUS) from Escherichia coli as a marker.
The DNA was injected into the soybeans using the particle acceleration method or gene gun.
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GENE CONSTRUCT
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introduction, herbicide-resistant soybeans have been quickly adopted. In 2010, 93% of all soybeans grown in the USA were herbicide-resistant, as were 78% of all cotton and 70% of all maize varieties (http://www.ers.usda.gov/Data/BiotechCrops/).
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Soybean Industry Portfolio Quality/Food
Agronomic
2020
Modified 7S Protein FF
(Pioneer/DuPont)
High-Oleic, Stearate
(Pioneer/DuPont)
Source: Pipeline from Industry Sources; prepared by ASA, USSEC, USB. Updated January, 2010
Nematode Resistance
(Monsanto; Syngenta;
Pioneer/DuPont)
LibertyLink (Bayer)
Imidazolinone TolerantBrazil only.
(BASF/Embrapa Brazil)RR2Y(Monsanto)
Dicamba Tolerant
(Monsanto)
Omega-3 Stearidonic Acid
(Monsanto)
High Beta- Conglycinin
(Pioneer/DuPont)
Low-Phytate(Pioneer/DuPont)
Feed: High Protein Soybean
(Pioneer/DuPont)
Omega-3EPA/DHA
(Pioneer/DuPont)
2010 2012
GAT/Glyphosate-ALS Soybean
(Pioneer/DuPont) Bt/RR2YBrazil only
(Monsanto)
HPPD Tolerant (Syngenta)
High Oleic / Low-Sat
(Monsanto)
HighStearate(Monsanto;
Pioneer/DuPont)
Pipeline of biotech events and novel trait releases
2,4-D Tolerant(Dow)
Disease Resistance
(Syngenta; Pioneer/DuPont)
Glufosinate & Isoxaflutole Tolerant (Bayer/MS Technologies)
Low Raff-Stach
(Virginia Tech)
RVSD Biotech Pipeline 02-04-10 V3
Commercialized
Glufosinate & Isoxaflutole Tolerant
& LibertyLink(Bayer/MS Technologies)
High-Oleic(Pioneer/DuPont)
Low-Linolenic(Syngenta)
Higher Yield I(Monsanto;
Pioneer/DuPont)
Higher Yield II(Monsanto;
Pioneer/DuPont)
Rust(Monsanto; Syngenta;
Pioneer/DuPont)
Aphid Resistance(Monsanto)
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Statistical data
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Grown in:Argentina, Australia, Canada, Chile, China, France, Germany, India, Mexico, South Africa, Spain, Uruguay, USA
Not yet grown?:Brazil, Egypt, other EU, Japan, Kenya, Korea, Switzerland
Traits:Herbicide-tolerant Insect resistantViral resistantMale sterile/restorersDelayed ripeningOil contentVitamin A, vaccines
GM-crops: 53 million ha (2001); 62.3% RR-soybean
Transgenic crops
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Regulation of Plant Biotechnology in the United States
The U.S. Coordinated Framework (1986)• The potential risks posed by genetically engineered
organisms are not fundamentally different from those posed by conventional products
• Regulation should be science-based and oversight conducted on a case-by-case basis
• Existing laws provide adequate authority
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Coordination of U.S. Agencies
FDA Safe for usefood and feed
USDASafe for agriculture and the environment
EPA Safe for usein pesticides
Products are regulated according to their intended use, with some products being regulated under more than one agency.
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Regulation under the Coordinated Framework
New Trait/Crop Agency ReviewInsect resistance in food crop(Bt corn)
USDAEPAFDA
Agricultural and environmental safety Environmental, food safety of pesticideFood/feed safety
Herbicide tolerance in food crop(Roundup Ready soybeans)
USDAEPAFDA
Agricultural and environmental safetyNew herbicide useFood/feed safety
Herbicide tolerance in ornamental crop
USDAEPA
Agricultural and environmental safetyNew herbicide use
Modified oil in food crop(High oleic acid soybeans)
USDAFDA
Agricultural and environmental safetyFood/feed safety
Modified flower color (Blue roses)
USDA Agricultural and environmental safety20
Regulation under the Coordinated Framework
• Corn - HT, IR, AP• Soybean - HT, PQ• Cotton - HT, IR• Canola - HT, AP, PQ• Papaya – VR• Squash – VR• Tobacco – PQ• Sugar beet - HT
Tomato - PQ Chicory – AP Potato - IR, VR Rice – HT Flax – AP Plum - VR
HT – herbicide toleranceIR – insect resistanceAP – agronomic propertiesVR – virus resistancePQ – product quality
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Transgenic crops: cotton, cucumber, melon, maize, tomato, papaya, potato, soybean, canola, sugar beet, tobacco, carnation
In pipeline: sweetpotato, cassava, banana/plantain, groundnut, chickpea, pigeonpea, pea, cowpea, sorghum, wheat
Transgenic technology: from test tubes tofarmers fields
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Case study#1Nature Biotechnology 22, 204 - 209 (2004) Published online: 18 January 2004 | doi:10.1038/nbt934Assessing the survival of transgenic plant DNA in the human gastrointestinal tractTrudy Netherwood1,2, Susana M Martín-Orúe1, Anthony G O'Donnell2, Sally Gockling1,2, Julia Graham1,2, John C Mathers3,4 & Harry J Gilbert1
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Introduction The inclusion of genetically modified (GM) plants in the human diet has raised concerns
about the possible transfer of transgenes from GM plants to intestinal microflora and
enterocytes. The persistence in the human gut of DNA from dietary GM plants is
unknown. Here they study the survival of the transgene epsps from GM soya in the
small intestine of human ileostomists (i.e., individuals in which the terminal ileum is
resected and digesta are diverted from the body via a stoma to a colostomy bag).
The amount of transgene that survived passage through the small bowel varied among
individuals, with a maximum of 3.7% recovered at the stoma of one individual. The
transgene did not survive passage through the intact gastrointestinal tract of human
GM soya. Three of seven ileostomists showesubjects fed d evidence of low-frequency
gene transfer from GM soya to the microflora of the small bowel before their
involvement in these experiments. 24
Methods rate of DNA degradation
DNA survival in humans with an intact gastrointestinal tract We fed the test meal containing GM soya to 12 human volunteers (with intact gastrointestinal tracts) and quantified the presence of the transgene in feces by PCR. For all volunteers, 90–98% of the indi-gestible marker was recovered in the feces but the transgene was not detected.
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Result
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DNA survival in humans with an intact gastrointestinal tract
They fed the test meal containing GM soya to 12 human volunteers
(with intact gastrointestinal tracts) and quantified the presence of the
transgene in feces by PCR. For all volunteers, 90–98% of the indi- gestible
marker was recovered in the feces but the transgene was not detected. This was
not the result of PCR inhibition because a 180 bp product was amplified by
PCR when the fecal material was spiked with 400 copies of the transgene in
the aliquot used in the PCRs (data not shown). Thus, although the epsps
transgene can survive passage through the small bowel of ileostomists, it is
completely degraded in the large intestine
Case study # 2
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Introduction Transgenic rice (Oryza sativa L.) containing two agronomically useful genes was
generated using Agrobacterium LBA4404 with an additional virulence plasmid,
virG(pTiBo542)/virE1virE2(pTiA6).The plants were transformed with
phosphinothricin acetyl transferase (pat) gene for herbicide resist-ance and
Bacillus thuringiensis (Bt) crystal insecticidal protein gene for insect resistance.
The herbicide application test of the progeny
from the three sets of primary plants showed that the transferred pat gene was stably
expressed in the T1generation. The insect feeding bioassay with T1generation plants
conferring resistance to her-bicide and established that the transgenic plants having a
complete Bt gene were toxic to tobacco bud-worm (Heliothis virescens) larvae
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Materials and MethodsMature seed-derived callus from tropical rice (Oryza sativa L. var. ja-ponica) cv. Maybelle was used for Agrobacterium-mediated transfor-mation. Bacterial strain and plasmids Agrobacterium strain, LBA4404 (octopine, Hoekema et al. 1983), wasused. The plasmid construct used was pAGM281 (provided by Myco-gen Plant Sciences) containing the ubi/pat/orf25\bt\(4Ocs)∆mas ex-pression vecto
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Results
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Results In summary, mature seed from a commercial rice cultivar,
Maybelle, was used to generate callus. Callus was trans-
formed using A. tumefaciens LBA4404 carrying two agronom-ically useful
genes, herbicide and insect resistance and the pCH32 helper plasmid with
virulence genesThis study demonstrates that a commercial rice cultivar
has been engineered with economically significant genes
conferring resistance to a herbicide and insect feeding using
Agrobacterium-mediated transformation.
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Controversies Violations of human rights as a result of GM soyWednesday, 23 November 2011 12:34GMOs in agriculture again a theme on UN-committee of human rights:Violations of Human Rights as a result of the Genetic-Modified Soya-Monocultures -- the Right to Food and Health - for the Argentinean Population, Farmers and Bee-KeepersGRR wrote report for UN-committee – Hearing in Geneva 14 November.on the consequences of a highly perturbing global development, the use of genetically modified GM-soya in Argentina. This seriously violates the economic, social and cultural rights of the Argentinean Population, Farmers and Bee-KeepersArgentina in particular is one of the most affected countries worldwide by this development.The GM agro-biotechnology accelerates the extinction of small farmers around the world. Deforestation, an increasing use of pesticides, destruction of livelihoods of indigenous peoples, small farmers and peasants, land concentration, slave labour, the rural exodus and increasing poverty in the countryside are the effects already abundantly demonstrated by the consequences of an agro-industrial model of transgenic monoculture.
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Nature Biotechnology | News Glyphosate resistance threatens Roundup hegemony
Nature Biotechnology28,537–538,(2010) doi:10.1038/nbt0610-537 Corrected online13 October 2010 These marestail plants infesting a crop of wheat in Tennessee
are resistant to glyphosate herbicide Roundup. Weeds are becoming increasingly resistant to glyphosate, a
report from the US National Academy of Sciences (NAS) released in April has found. The driving force, according to the report, is farmers' dependence on the weed killer accompanied by the widespread adoption of genetically modified (GM) herbicide-tolerant crops. Seed makers are hoping to forestall the problem by developing GM crops with 'stacked' traits that tolerate multiple herbicides. But weed scientists warn that if farmers manage these new crops in the same way as they managed their glyphosate-tolerant predecessors, weeds will simply become resistant to the new technologies.
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SOYBEAN TRANSGENIC EVENT MON87705 AND METHODS FOR DETECTION THEREOF1977/CHENP/2011 A (1977/CHENP/2011)Filed on 2011-03-21Publication date 2011-12-02
The present invention provides a transgenic soybean event MON87705, and cells, seeds, and plants comprising DNA diagnostic for the soybean event. The invention also provides compositions comprising nucleotide sequences that are diagnostic for said soybean event in a sample, methods for detecting the presence of said soybean event nucleotide sequences in a sample, probes and primers for use in detecting nucleotide sequences that are diagnostic for the presence of said soybean event in a sample, growing the seeds ofsuch soybean event into soybean plants, and breeding to produce soybean plants comprising DNA diagnostic for the soybean event.
ApplicantMONSANTO TECHNOLOGY LLCMAIL ZONE E1NA, 800 N. LINDBERGH BLVD., ST. LOUIS, MO 63167 U.S.A.
InventorWAGNER, NICHOLAS, BURNS, WEN C., GODSY, ERIC J., ROBERTS, PETER D.
International InformationClassificationA01H1/02Publication numberWO 2010/037016 A1Application date2009-09-28Application numberPCT/US2009/058591
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Conclusion GM crops gives protection from biotic factors like insect,
pathogen as well as abiotic factors like drought, salinity, flood
etc.
In advance agriculture herbicide resistant GM crops plays vital
role.
Now a days two agronomically important genes transfer into
single plant leads to increase potential of the crops.
It really helps in growth and development of agriculture sector.
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References Albert l. Lehninger., biochemistry, second edition, pg no. 708-
709. R.C.Dubey., A text book of biotechnology, fourth edition, pg
no.323-327. Nature Biotechnology 22, 204 - 209 (2004)
Published online: 18 January 2004 | doi:10.1038/nbt934 P lant P hysiol. 158. 1221–1226(2001) Urban&F
ischerVerlag http://www.urbanfischer.de/journals/jpp International Journal of Molecular Sciences ISSN 1422-0067 www.mdpi.com/journal/ijm
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Thank you
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