biotechnological approaches in Weed management.ppt

biotechnological approaches in Weed management

1. Biotechnological Biotechnological
approaches for weed approaches for weed
managementmanagement

2. 
Weed – 5 to 100 % loss Weed – 5 to 100 % loss

Due to – rare fallowing, much Due to – rare fallowing, much
monoculturemonoculture

Highly intensive agricultureHighly intensive agriculture

High yield index crop (wimpish High yield index crop (wimpish
competitiveness with weeds)competitiveness with weeds)

3. 
Herbicides – most effective Herbicides – most effective

Nonselective Nonselective

Resistance development in weedsResistance development in weeds

Crop specific weeds Crop specific weeds
Legume weeds in soybeanLegume weeds in soybean
Abutilon indicum Abutilon indicum in cottonin cotton
Bromes & barleys in wheatBromes & barleys in wheat
EchinoclovaEchinoclova in rice in rice

8. How biotech can help in weed control?How biotech can help in weed control?

Direct Direct

by increasing crops competitiveness with by increasing crops competitiveness with
weeds using exogenous genesweeds using exogenous genes

By genetically engineering crops wuth By genetically engineering crops wuth
genes conferring resistance to herbicidesgenes conferring resistance to herbicides

Biotechnologically cultivating and possibly Biotechnologically cultivating and possibly
modifying biocontrol organisms.modifying biocontrol organisms.

9. How biotech can help in weed control?How biotech can help in weed control?

Indirect Indirect

Genes for producing natural allelochemicals Genes for producing natural allelochemicals
(phytoalexins, Strychnine, Nicotine) Safety?(phytoalexins, Strychnine, Nicotine) Safety?

Genes for enhanced nutrient uptake (N, P, Fe) Use Genes for enhanced nutrient uptake (N, P, Fe) Use
of microbial genes with root specific promotorsof microbial genes with root specific promotors

Increased growth rate (Smothering effect, leaf Increased growth rate (Smothering effect, leaf
morphologies, angles - quickly shade out weed) – morphologies, angles - quickly shade out weed) –
Yield index?Yield index?

Add genes for pathogen or arthropod resistance to Add genes for pathogen or arthropod resistance to
enhance selectivity by biocontrol agentsenhance selectivity by biocontrol agents

Long term approachesLong term approaches

10. Direct approachesDirect approaches

Adding genes for herbicide toleranceAdding genes for herbicide tolerance

11
stst
achievement in plant genetic achievement in plant genetic
engineeringengineering

Glyphosate, glufosinate (Bialaphos, Glyphosate, glufosinate (Bialaphos,
phosphinothricin), sulfonyl urea etc. phosphinothricin), sulfonyl urea etc.

11. Genes for herbicide resistanceGenes for herbicide resistance

aroAaroA from from AgrobacteriumAgrobacterium sp. Strain CP4 sp. Strain CP4

Glyphosate/ RoundupGlyphosate/ Roundup

Encodes glyphosate insensitive EPSPS Encodes glyphosate insensitive EPSPS
involved in amino acid biosynthesis involved in amino acid biosynthesis
pathwayspathways

5-enol pyruvyl shikimate 3- phosphate synthase 5-enol pyruvyl shikimate 3- phosphate synthase

12. Genes for herbicide resistanceGenes for herbicide resistance

bar bar from from Streptomyces hygroscopicusStreptomyces hygroscopicus

Bialaphos /Basta/ GlufosinateBialaphos /Basta/ Glufosinate

Encodes for the enzyme Encodes for the enzyme
Phosphinothricin acetyl transferase Phosphinothricin acetyl transferase
involved in acetylating phosphinothricininvolved in acetylating phosphinothricin

13. Genes for herbicide resistanceGenes for herbicide resistance

bxn bxn from from Klebsiella ozaeneKlebsiella ozaene

BromoxynilBromoxynil

Encodes for the enzyme Bromoxynil nitrilaseEncodes for the enzyme Bromoxynil nitrilase
Toxic herbicide Toxic herbicide
Bromoxynil nitrilaseBromoxynil nitrilase
3,5 dibromo -4-hydroxy benzoic acid3,5 dibromo -4-hydroxy benzoic acid

14. Genes for herbicide resistanceGenes for herbicide resistance

gox gox from from Achromobacter Achromobacter sp strain LBAAsp strain LBAA

Glyphosate/ round upGlyphosate/ round up

Encodes for the enzyme glyphosate oxido reductaseEncodes for the enzyme glyphosate oxido reductase
Toxic Glyphosate Toxic Glyphosate

Glyphosate oxido reductaseGlyphosate oxido reductase
Amino methyl phosphonic acid + Glyoxylate (non-Amino methyl phosphonic acid + Glyoxylate (non-
toxic)toxic)

15. Genes for herbicide resistanceGenes for herbicide resistance

Hra, csr I-1, c3 and ahas 3r genes Hra, csr I-1, c3 and ahas 3r genes

From From Arabidopsis thaliana, N. tabacum, yeast & Arabidopsis thaliana, N. tabacum, yeast &
E. coliE. coli

Sulfonyl ureas’ and ImidazolononesSulfonyl ureas’ and Imidazolonones

Encodes for the enzyme ALS (Insensitive)Encodes for the enzyme ALS (Insensitive)

Aceto lactate synthase (involved in amino acid Aceto lactate synthase (involved in amino acid
biosynthetic pathway)biosynthetic pathway)

16. Targets of HerbicidesTargets of Herbicides
InhibitorsInhibitors

ALS- Aceto lactate synthase ALS- Aceto lactate synthase

ACCase- Acetyl CoA CorboxylaseACCase- Acetyl CoA Corboxylase

PS II- Photo system II PS II- Photo system II

17. Glyphosate actionGlyphosate action

Broad spectrum herbicideBroad spectrum herbicide

Inhibits- EPSPS Inhibits- EPSPS
Aromatic amino acid biosynthesis in plantsAromatic amino acid biosynthesis in plants
Starving the cells of aromatic amino acidStarving the cells of aromatic amino acid
Disruption of protein synthesisDisruption of protein synthesis
Killing of plantsKilling of plants

20. Strategies for glyphosate resistanceStrategies for glyphosate resistance
Transgenes for any one of the following Transgenes for any one of the following
purposespurposes
i.i.Over production of EPSPS enzymeOver production of EPSPS enzyme
ii.ii.Production of insensitive/ tolerant EPSPS Production of insensitive/ tolerant EPSPS
enzymeenzyme
iii.iii.Production of an enzyme that inactivates Production of an enzyme that inactivates
glyphosateglyphosate
iv.iv.Combination of ii and iiiCombination of ii and iii

21. EPSPS
EPSPS
Proteins
CO
2
H
2
O
NH
3
aromatic
amino acids
Proteins
CO
2
H
2
O
NH
3
aromatic
amino acids
glyphosate
Proteins
CO
2
H
2
O
NH
3
aromatic
amino acids
glyphosate
CP4 EPSPS
EPSPS

22. Strategies for glyphosate resistanceStrategies for glyphosate resistance
i. Over production of EPSPS enzymei. Over production of EPSPS enzyme
Petunia hybrida Petunia hybrida EPSPS – gene isolated and over EPSPS – gene isolated and over
expressed in other plants.expressed in other plants.
Glyphosate tolerance incorporatedGlyphosate tolerance incorporated
Under field condition insufficient protectionUnder field condition insufficient protection

23. Strategies for glyphosate resistanceStrategies for glyphosate resistance
ii. Production of insensitive/ tolerant EPSPS enzymeii. Production of insensitive/ tolerant EPSPS enzyme
Mutants of Mutants of Salmonella typhimuriumSalmonella typhimurium and and E. coli E. coli produced produced
mutant form of EPSPS tolerant to glyphosatemutant form of EPSPS tolerant to glyphosate
Expressed in Tobacco, soybean, Canola and tomato – Expressed in Tobacco, soybean, Canola and tomato –
Inadequate for commercial use.Inadequate for commercial use.
AgrobacteriumAgrobacterium CP4 strain- Tolerant EPSPS CP4 strain- Tolerant EPSPS
Introduced to Soybean- No visible herbicide injury in Introduced to Soybean- No visible herbicide injury in
fieldfield
Then introduced to cotton & other cropsThen introduced to cotton & other crops

24. Strategies for glyphosate resistanceStrategies for glyphosate resistance
iii. Production of an enzyme that inactivates iii. Production of an enzyme that inactivates
glyphosateglyphosate

gox gox from from Achromobacter Achromobacter sp strain LBAAsp strain LBAA
Glyphosate Glyphosate
Glyphosate oxido reductaseGlyphosate oxido reductase
Amino methyl Amino methyl
phosphonic acidphosphonic acid + + Glyoxylate (non-toxic)Glyoxylate (non-toxic)
Expression of Expression of gox gox and localisation in to chloroplastand localisation in to chloroplast
Excellent vegetative and reproductive tolerance Excellent vegetative and reproductive tolerance

25. Strategies for glyphosate resistanceStrategies for glyphosate resistance
iv. Combination of ii and iiiiv. Combination of ii and iii
goxgox gene + gene + AgrobacteriumAgrobacterium sp CP4 EPSPS gene sp CP4 EPSPS gene
Eg. Roundup Ready Maize (Monsanto, USA)Eg. Roundup Ready Maize (Monsanto, USA)

26. Cases of resistance to glyphosate & Cases of resistance to glyphosate &
their mechanismtheir mechanism
OrganismOrganism SourceSource MechanismMechanism
1.Tobacco1.Tobacco Cell lines selected for Cell lines selected for
resistanceresistance
Overproduction of Overproduction of
EPSPS by gene EPSPS by gene
amplificationamplification
2. Carrot2. Carrot Cell lines selected for Cell lines selected for
resistanceresistance
Overproduction of Overproduction of
EPSPS by gene EPSPS by gene
amplificationamplification
3. 3. Salmonella Salmonella
typhimuriumtyphimurium
Mutant strainMutant strain Tolerant EPSPSTolerant EPSPS
(Pro at position 101 (Pro at position 101
replaced by Ser)replaced by Ser)
4.4. E. coli E. coli Mutant strainMutant strain (Ala at position 96 (Ala at position 96
replaced by Gly)replaced by Gly)
5. 5.
AgrobacteriumAgrobacterium
Mutant strain CP4Mutant strain CP4 High tol. of EPSPS and High tol. of EPSPS and
binding to PEPbinding to PEP

27. Cases of resistance to glyphosate & Cases of resistance to glyphosate &
their mechanismtheir mechanism
OrganismOrganism SourceSource MechanismMechanism
6. 6. E. coliE. coli Site directed mutagenesisSite directed mutagenesisTol. of EPSPS due to Tol. of EPSPS due to
amino acid amino acid
replacementreplacement
7. 7. A. thalianaA. thaliana Site directed mutagenesisSite directed mutagenesisTol. of EPSPS due to Tol. of EPSPS due to
amino acid amino acid
replacementreplacement
8. 8. PetuniaPetunia Site directed mutagenesisSite directed mutagenesisTol. of EPSPS due to Tol. of EPSPS due to
amino acid amino acid
replacementreplacement
9.9. Achromobacter Achromobacter
sp. Strain LBAAsp. Strain LBAA
Natural isolate Natural isolate Inactivation by Inactivation by
glyphosate glyphosate
oxidoreductase oxidoreductase

28. Class of herbicide Compound/herbicideTransgene/
mechanism CompaniesCrops
Glycine Glyphosate (Roundup)' Agrobacterium CP4
‑
Monsanto Soybean, rape, tomato
resistant gene
Glyphosate (Roundup) Maize resistant gene Monsanto Maize
Glyphosate (Roundup) Oxidoreductase Monsanto Maize, rape, soybean
Detoxification
Phosphinic acid Phosphinothricin bar gene PAT
‑
Hoechst/AgrEvo/ Maize, rice, wheat,
(Basta), () phosphinothricin Aventis Cotton, rape, potato,
acetyltransferase Novartis/Syngenta tomato, sugar beet
detoxification
Sulphonylurea Chlorsulphuron Mutant plant DuPont
‑
Rape, rice, flax, tomato,
(Glean) acetolactate synthase Pioneer Hi Bred
‑
sugar beet, maize
S triazines
‑
Atrazine (Lasso) Mutant plant DuPont, Ciba
‑
Soybean
chloroplast psbA Geigy/Novartis
gene
Nitriles Bromoxynil (Buctril) Nitrilase Calgene Cotton, rape, potato,
detoxification tomato
Phenoxy
‑
2,4 D
‑
Monooxygenase Schering/AgrEvo Maize, cotton
carboxylic acids detoxification

29. Class of herbicide
Compound/herbicide
Transgene/ mechanism
Companies
Crops
Glycine Glyphosate (Roundup)'Agrobacterium CP4‑ Monsanto Soybean, rape, tomato
resistant gene
Glyphosate (Roundup)Maize resistant geneMonsanto Maize
Glyphosate (Roundup)Oxidoreductase Monsanto Maize, rape, soybean
Detoxification
Phosphinic acid Phosphinothricin bar gene‑ Hoechst/AgrEvo/ Maize, rice, wheat,
(Basta), () phosphinothricin Aventis Cotton, rape, potato,
acetyltransferase Novartis/Syngentatomato, sugar beet
detoxification
Sulphonylurea Chlorsulphuron Mutant plant DuPont‑ Rape, rice, flax, tomato,
(Glean) acetolactate synthasePioneer Hi‑Bred sugar beet, maize
S‑triazines Atrazine (Lasso) Mutant plant DuPont, Ciba‑ Soybean
chloroplast psbA Geigy/Novartis
gene
Nitriles Bromoxynil (Buctril)Nitrilase Calgene Cotton, rape, potato,
detoxification tomato
Phenoxy‑ 2,4‑D Monooxygenase Schering/AgrEvo Maize, cotton
carboxylic acids detoxification
Herbicide‑resistant crops now in the field

30. Gene ConstructGene Construct

The gene is introduced into the plant The gene is introduced into the plant
nuclear genome via different nuclear genome via different
mechanismsmechanisms

The trait is then incorporated to elite The trait is then incorporated to elite
varieties by backcrossing varieties by backcrossing
promoter (leading sequence) resistant EPSPS terminator

32. TGGAAAAGGAAGGTGGCTCCTACAAATGCCATCATTGCGATAAAGGAAAGGCCATCGTTGAAGATGCCTCTGCCGACAGTGGTCCCAAAG
ATGGACCCCCACCCACGAGGAGCATCGTGGAAAAAGAAGACGTTCCAACCACGTCTTCAAAGCAAGTGGATTGATGTGATATCTCCACTGA
CGTAAGGGATGACGCACAATCCCACTATCCTTCGCAAGACCCTTCCTCTATATAAGGAAGTTCATTTCATTTGGAGAGGACACG CTGACAAG
CTGACTCTAGCAGATCTTTCAAGAATGGCACAAATTAACAACATGGCACAAGGGATACAAACCCTTAATCCCAATTCCAA TTTCCATAAACC
CCAAGTTCCTAAATCTTCAAGTTTTCTTGTTTTTGGATCTAAAAAACTGAAAAATTCAGCAAATTCTATGTTGGTTTTGAAAAAAGATTCAATT
TTTATGCAAAAGTTTTGTTCCTTTAGGATTTCAGCATCAGTGGCTACAGCCTGCATGCTTCACGGTGCAAGCAGCCGGCCCGCAACCGCCCG
CAAATCCTCTGGCCTTTCCGGAACCGTCCGCATTCCCGGCGACAAGTCGATCTCCCACCGGTCCTTCATGTTCGGCGGTCTCGCGAGCGGTG
AAACGCGCATCACCGGCCTTCTGGAAGGCGAGGACGTCATCAATACGGGCAAGGCCATGCAGGCCATGGGCGCCAGGATCCGTAAGGAA
GGCGACACCTGGATCATCGATGGCGTCGGCAATGGCGGCCTCCTGGCGCCTGAGGCGCCGCTCGATTTCGGCAATGCCGCCACGGGCTGC
CGCCTGACCATGGGCCTCGTCGGGGTCTACGATTTCGACAGCACCTTCATCGGCGACGCCTCGCTCACAAAGCGCCCGATGGGCCGCGTGT
TGAACCCGCTGCGCGAAATGGGCGTGCAGGTGAAATCGGAAGACGGTGACCGTCTTCCCGTTACCTTGCGCGGGCCGA AGACGCCGACGC
CGATCACCTACCGCGTGCCGATGGCCTCCGCACAGGTGAAGTCCGCCGTGCTGCTCGCCGGCCTCAACACGCCCGGCATCACGACGGTCAT
CGAGCCGATCATGACGTGCGATCATACGGAAAAGATGCTGCAGGGCTTTGGCGCCAACCTTACCGTCGAGACGGATGCGGACGGCGTGC
GCACCATCCGCCTGGAAGGCCGCGGCAAGCTCACCGGC CAAGTCATCGACGTGCCGGGCGACCCGTCCTCGACGGCCTTCCCGCTGGTTG
CGGCCCTGCTTGTTCCGGGCTCCGACGTCACCATCCTCAACGTGCTGATGAACCCCACCCGCACCGGCCTCATCCTGACGCTGCAGGAAATG
GGCGCCGACATCGAAGTCATCAACCTGCGCCTTGCCGGCGGCGAAGACGTGGCGGACCTGCGCGTTCGCTCCTCCACGCTGAAGGGCGTC
ACGGTGCCGGAAGACCGCGCGCCTCCGATGATCGACGAATATCCGATT CTCGCTGTCGCCGCCGCCTTCGCGGAAGGGGCGACCGTGATG
AACGGTCTGGAAGAACTCCGCGTCAAGGAAAGCGACCGCCTCTCGGCCGTCGCCAATGGCCTCAAGCTCAATGGCGTGGATTGCGATGAG
GGCGAGACGTCGCTCGTCGTGCGTGGCCGCCCTGACGGCAAGGGGCTCGGCAACGCCTCGGGCGCCGCCGTCGCCACCCATCTCGATCAC
CGCATCGCCATGAGCTTCCTCGTCATGGGCCTCGTGTCGGAAAACCCTGTCACGGTGGACGATGCCACGATGATCGCCACGAGCTTCCCGG
AGTTCATGGACCTGATGGCCGGGCTGGGCGCGAAGATCGAACTCTCCGATACGAAGGCTGCC TGATGAGCTCGAATTCGAGCTCGGTACC
GGATCCAATTCCCGATCGTTCAAACATTTGGCAATAAAGTTTCTTAAGATTGAATCCTGTTGCCGGTCTTGCGATGATTATCATATAATTTCT
GTTGAATTACGTTAAGCATGTAATAATTAACATGTAATGCATGACGTTATTTATGAGATGGGTTTTTATGATTAGAGTCCCGCAATTATACAT
TTAATACGCGATAGAAAACAAAATATAGCGCGCAAACTAGGATAAATTATCGCGCGCGGTGTCATCTATGTTACTAGATCGGGGATCGATC
CCCCACCGGTCCTTCATGTTCGGCGGTCTCGCGAGCGGTGAAACGCGCATCACCGGCCTTCTGGAAGGCGAGGACGTCATCAATACGG GC
AAGGCCATGCAGGCCATGGGCGCCAGGATCCGTAAGGAAGGCGACACCTGGATCATCGATGGCGTCGGCAATGGCGGCCTCCTGGCGCC
TGAGGCGCCGCTCGATTTCGGCAATGCCGCCACGGGCTGCCGCCTGACCATGGGCCTCGTCGGGGTCTACGATTTCAAGCGCATCATGCTG
GGAA

Gene Construct: RR soybeanGene Construct: RR soybean

LOCUS AB209952.1LOCUS AB209952.1

Size 2457 bpSize 2457 bp

SOURCE Glycine maxSOURCE Glycine max

FEATURES FEATURES

CaMV35SCaMV35S

Chloroplast transit Chloroplast transit
peptidepeptide

CP4EPSPS CP4EPSPS

Nos 3’ terminatorNos 3’ terminator

Repeated fragment Repeated fragment
of cp4epsps of cp4epsps
truncatetruncate
Gene bank gi|62318478|dbj|AB209952.1|[62318478]