biotechnology gm technology for herbicide
TRANSCRIPT
Biotechnology
Plant biotechnology and crop improvement
GM technology for Herbicide
Paper No. : 12 Plant biotechnology Biotechnology and crop improvement
Module : 34 GM technology for Herbicide
Principal Investigator: Dr Vibha Dhawan, Distinguished Fellow and Sr. Director The Energy and Resources Institute (TERI), New Delhi
Co-Principal Investigator: Prof S K Jain, Professor, of Medical Biochemistry Jamia Hamdard University, New Delhi
Paper Coordinator: Dr Vibha Dhawan, Distinguished Fellow and Sr. Director The Energy and Resources Institute (TERI), New Delhi
Content Writer: Dr Gulshan Chaudhary, Post Doctorate Fellow, The Energy and Resources Institute (TERI), New Delhi
Content Reviewer: Dr Vibha Dhawan, Distinguished Fellow and Sr. Director
The Energy and Resources Institute (TERI), New Delhi
Biotechnology
Plant biotechnology and crop improvement
GM technology for Herbicide
Description of Module
Subject Name Biotechnology
Paper Name Plant biotechnology and crop improvement
Module Name/Title GM technology for Herbicide
Module Id 34
Pre-requisites
Objectives
Keywords
Biotechnology
Plant biotechnology and crop improvement
GM technology for Herbicide
GM TECHNOLOGY FOR HERBICIDES
TABLE OF CONTENTS
1. Learning Objectives
2. Introduction
2.1 Why develop herbicide resistance crops?
3. Glyphosate Resistance
3.1 EPSPSs insensitive to glyphosate
3.2 Detoxification of glyphosate
4. Glufosinate Resistance
4.1 Detoxification of glufosinate molecule
5. Safety aspects of herbicide tolerant technology
5.1 Toxicity and Allergenicity
5.2 Effects on the Plants
5.3 Persistence or invasiveness of crops
6. Advantages of Herbicide Tolerant Crops
7. Disadvantages of transgenic herbicide resistance
8. Current status of herbicide tolerance
9. Summary
10. Figures and Table
Biotechnology
Plant biotechnology and crop improvement
GM technology for Herbicide
1. LEARNING OBJECTIVES
In the present module, we aim GM crops, Herbicide tolerance crops, its mode of action.
Additionally, we give a preview to understand past, present and future scenario of herbicide
tolerant plants.
2. INTRODUCTION
Weeds cause loss of crop production globally every year. The chemical herbicides which are
used is harmful as, some time it causes harm to plants also. So to overcome chemical
herbicides side effect now days genetic engineering is used to incorporate herbicides
resistance gene in crops know as genetically modified crops (GM).
GM for Herbicide can be done by two type: (1) Target site and, (2) Non target site herbicide
resistance.
1. Target site: In this the target site of herbicide is changed like mutation in amino acid so that
the herbicide does not binds to the site and its effect is lethal to plant.
2. Non target site: In this changes is done in such a way that the amount of herbicide reaching
the target site is very low(non lethal).
2.1 Why develop herbicide resistance crops?
Different herbicides have different Mode of Actions (MOAs) which mean that a target
enzyme no longer functions properly, or at all. This is usually because the molecule of
herbicide has distorted the enzyme molecule. Enzymes are also known to be catalysts
and they provides a platform for specific chemical reaction in which they act directly
or indirectly. The Knocking-out’ of an enzyme with a herbicide shows two main effects
with various consequences:
Chemicals components of the reaction accumulate by damaging directly or indirectly.
Absence of the reaction’s product will restrict growth, either through starvation of
key building blocks or because the reaction makes chemicals which normally protect
the plant.
The enzyme functioning normally Figure 1 (1) while in figure 1(2) molecule of herbicide
stopping biochemically by coming together to form the usual product. As a result the plant
dies due to the damage of building locks. While in figure 1 (3) A mutated enzyme combines
with the herbicide to continue the reaction. Either herbicide is selective to species with this
Biotechnology
Plant biotechnology and crop improvement
GM technology for Herbicide
type of enzyme, or an individual plant with this mutation could reproduce to establish a
population of resistant weeds.
Examples of gene-based herbicide resistance
Herbicide(s) Effected part/function Mode of development of
herbicide resistance
GM plant
Triazines Photosynthesis
Resistance is due to an
alteration in the psbA
gene, which codes for the
target of this herbicide,
chloroplast protein D-1.
Canola
Sulfonylureas
Inhibitors of AHAS
therefore stopping the
synthesis of branched-
chain amino acids,with
subsequent plant death.
Genes encoding resistant
versions of the enzyme
acetolactate synthetase.
Soybean
,Poplar,
Canola, Flax,
and Rice
Imidazolinones
Inhibitors of AHAS by
stopping synthesis of
the branched-chain
amino acids, with
subsequent plant death.
Strains with resistant
versions of the enzyme
acetolactate synthetase
have been selected in
tissue culture.
Maize, Canola,
Wheat and
Rice.
Aryloxphenoxypropionates,
Cyclohexanediones lipid biosynthesis
These herbicides inhibit
the enzyme acetyl
coenzyme
A carboxylase. Resistance,
selected in tissue culture,
is due either to an altered
enzyme that is not
herbicide sensitive or to
the degradation of the
herbicide.
Maize,
Soyabean and
Cotton.
Glyphosate
inhibition of EPSPS
activity which, disrupts
the shikimate pathway
and inhibits aromatic
amino acid production,
which ultimately
causing the death of
plant.
Resistance is from
overproduction of EPSPS,
the target of this herbicide.
Soybean,
Canola
Cotton, Maize
Bromoxynil
Inhibition of
acetolactate synthase
(ALS)
Resistance to this
photosystem II inhibitor
has
been created with a
bacterial nitrilase gene,
which encodes an enzyme
that degrades this
herbicide.
Tobacco,
Cotton and
Canola
Phenoxycarboxylic acids
(e.g., 2,4-D and 2,4,5-T)
Remain in soil and
toxic to lower mammals
and humans.
Transformation with the
tfdA gene from
Alcaligenes, which
Cotton and
tobacco
Biotechnology
Plant biotechnology and crop improvement
GM technology for Herbicide
encodes a dioxygenase
that degrades this
herbicide.
Glufosinate
(phosphinothricin)
Inhibits glutamine
Synthetase.
Over 20 different plants
have been transformed
with either the bar gene
from Streptomyces
hygroscopicus or the pat
gene from
S. viridochromogenes. The
phosphinothricin
acetyltransferase that these
genes encode
detoxifies this herbicide.
Canola, Corn,
Cotton, Maize,
Rice and
sugarbeet.
Cyanamide Inhibit catalase,
cytochromoxidase
Cyananide hydratase gene
from the fungus
Myrothecium verrucaria
was introduced. The
enzyme encoded by this
gene converts cyanamide
to urea.
Tobacco,
wheat and
soyabean.
3. Glyphosate Resistance:
It’s a very important herbicides used worldwide. Glyphosate inhibits an enzyme 5-
enolpyruvylshikimate-3-phosphate synthase (EPSPS) of shikimate pathway. An enzyme
enolpyruvylshikimate-3-phosphate synthase catalases the reaction by phosphoenolpyruvate
(PEP) and shikimate-3-phophate (S3P) to form a compound 5-enolpyruvylshikimate-3-
phosphate (EPSP), resulted in to the synthesis of aromatic amino acids. Diagram is showing
the shikimate pathway and glyphosate inhibiting pathway (fig.2).
Work to develop glyphosate resistant crops by genetic engineering are focused on
following strategies (Fig 3):
Overproduction of EPSP synthase.
Introduction of a metabolic detoxification gene.
Introduction of an altered EPSP synthase enzyme with decreased affinity for
glyphosate.
3.1 EPSPSs insensitive to glyphosate:
Since 1985 many researchers have identified many genes from different sources with same or
different mode of action which help plants combat glyphosate (Comai et al. 1985; Stalker et
al. 1985; Kishore et al. 1986; Padgette et al. 1991; Berry et al. 1992; Zhou et al. 1995, 2006;
Ye et al. 2001; Kahrizi et al. 2007). but CP4-EPSPS gene which is isolated from
Biotechnology
Plant biotechnology and crop improvement
GM technology for Herbicide
Agrobacterium strain. CP4, is best suited for transformation as it is insensitive to glyphosate.
CP4-EPSPS and sensitive EPSPS have identical binding site for substrate glyphosate. CP4-
EPSPS have high affinity for PEP then glyphosate, which allow the shikimate pathway to
function normally, as it ‘bypass’ the endogenous EPSPS (fig.3).
3.2 Detoxification of glyphosate:
Detoxification is another method which can be employed to confer glyphosate. Soil
microorganisms can metabolize glyphosate by two different ways (Fig. 4A): (a) cleavage of
the carbon–phosphorus bond, which results in the formation of phosphate and sarcosine (the
C-P lyase pathway)and (b) oxidative cleavage of the carbon–nitrogen bond on the carboxyl
side, catalyzed by glyphosate oxidoreductase (GOX), which results in the formation of
aminomethylphosphonic acid (AMPA) and glyoxylate (the AMPA pathway).
4. Glufosinate Resistance:
Glufosinate herbicides contain the active ingredient phosphinothricin, which kills plants by
blocking the enzyme responsible for nitrogen metabolism and for detoxifying ammonia, a by-
product of plant metabolism. Glutamine synthase (GS) catalyzes conversion of L-glutamate to
L-glutamine while assimilating ammonia in plants. Phosphinothricin (glufosinate, PPT), is a
herbicide that is analog of glutamate, inhibits glutamine synthase (GS) as L-glufosinate and
L-glutamate are very similar in structure, which results in accumulation of ammonium.
Futher research shows that PPT inhibits part of photorespiration process. Inhibition of GS
result in decrease in glutamate concentration which affect the photorespiration process by
accumulating glyoxylate that inhibits the RuBP-carboxylase that is CO2 formation. Crops
modified to tolerate glufosinate contain a bacterial gene that produces an enzyme that
detoxifies phosphinothricin and prevents it from doing damage.
4.1 Detoxification of glufosinate molecule:
Komoba and Sandermann, 1992 have shown that plants have endogenous metabolism of
glufosinate but its too slow to degrade the herbicide. So, gene from outside need to be
inserted to encode an enzyme that can detoxify L-glufosinate fast and prevent the herbicide
from reaching the target enzyme. Rasche, 1995; Vasil, 1996; Wehrmann et al., 1996 showed
two gene bar from Streptomyces hygroscopicu and pat gene from S. viridochromogenes
encodes Phosphinothricin N-Acetyltransferase (PAT) enzyme. PAT reduces L-glufosinate to
non-phytotoxic metabolite N-acetyl-L-glufosinate (NAG) which will prevent glufosinate to
inhibit GS (Fig.5). So, bar and pat gene are being used to transformation for GM plants
resistant to Glufosinate.
Biotechnology
Plant biotechnology and crop improvement
GM technology for Herbicide
5. Safety aspects of herbicide tolerant technology
5.1 Toxicity and Allergenicity:
In several countries agencies linked to Government regulatory have ruled that the crops
possessing herbicide tolerant conferring proteins and it do not poses any other environmental
and health risks as compared to non-GM counterparts. Proteins which are introduced are
assessed for the potential toxic and allergenic activity, according to the guidelines developed
by international organizations.
5.2 Effects on the Plants:
The main role of this protein that it expression did not harm the plant growth nor resulted
into the poor agronomic performance compared to parental crops, except for the
expression of an additional enzyme for herbicide tolerance or alteration of an already
existing enzyme, no other metabolic changes occur in the plant.
5.3 Persistence or invasiveness of crops
A major environmental concern associated with herbicide tolerant crops is their potential
to create new weeds through outcrossing with wild relatives or simply by persisting in the
wild themselves. This potential, however, is assessed prior to introduction and is also
monitored after the crop is planted. The current scientific evidence indicates that, in the
absence of herbicide applications, GM herbicide-tolerant crops are no more likely to be
invasive in agricultural fields or in natural habitats than their non-GM counterparts (Dale
et al., 2002).
The herbicide tolerant crops currently in the market show little evidence of enhanced
persistence or invasiveness.
.
6. Advantages of Herbicide Tolerant Crops
It is an excellent weed control, hence higher crop yields
Its flexibility because of its possibility to control the weeds later in the plant’s growth
It also reduces the numbers of sprays in a season therefore reduction in the fuel use
It’s also reduces soil compaction because of less need to go on the land to spray
Because of low toxicity compounds it do not remain active in the soil
It has ability to use conservation-till systems, with has consequent benefits to soil
structure and organisms (Felsot, 2000).
7. Disadvantages of transgenic herbicide resistance
Low toxicity to mammalian
Biotechnology
Plant biotechnology and crop improvement
GM technology for Herbicide
Its ecotoxicity as have side effects on soil microorganisms and agricultural flora ond
fauna
It develops some herbicide resistant weeds
Have some effect on yield performance
Having single selection pressure and weed resistance
Due to mutation it cause gene escape
It can also responsible for gene flow and contamination of organic crops
8. Current status of herbicide tolerance
Since 1996 to 2016, herbicide tolerat crops consistently occupying the largest planting area of
biotech crops. According to ISAAA in 2016, Herbicide Tolerant crops occupied 86.5 million
hectares or 47% of the 185.1 million hectares of biotech crops planted globally. According to
survey the most common are the glyphosate and glufosinate tolerant varieties. The table 1
shows countries that have approved major Herbicide Tolerant (with single and stacked genes)
crops for food, feed, and/or cultivation
9. Summary:
As we are evolving our techniques are evolving too. Conventional way of removing weed by
use chemical herbicides are not only harmful to weeds but it also effected environment, soil
texture and crop plants too. Herbicides resistance plants is better alternatives to conventional
techniques. When it come to technology then there is always an advantage as well as
disadvantage. So, we should be very careful. From 1996 to 2016, HT crops consistently
occupied the largest planting area of biotech crops. In 2016 alone, HT crops occupied 86.5
million hectares or 47% of the 185.1 million hectares of biotech crops planted globally. The
most common are the glyphosate and glufosinate tolerant varieties.
At present we can get many herbicides resistance plants in market. Still research is go on to
make most of the crop plants herbicides resistance.
Figures
Biotechnology
Plant biotechnology and crop improvement
GM technology for Herbicide
Figure 1-3Different modes of enzyme reaction
Biotechnology
Plant biotechnology and crop improvement
GM technology for Herbicide
Figure 2 Shikimate pathway that leads to the biosynthesis of aromatic amino acids and
mode of action of glyphosate on the reaction catalyzed by EPSPS.
Biotechnology
Plant biotechnology and crop improvement
GM technology for Herbicide
Figure 2 Strategy for the development of glyphosate-resistant
Table 1 Global adoption rate (%) for principal Biotech crops