Final report mst 5
-
Upload
hiccups-demafeliz -
Category
Documents
-
view
393 -
download
0
Transcript of Final report mst 5
![Page 1: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/1.jpg)
1
Principles and Practice of Environmental Safety Assessment
of Transgenic Plants
![Page 2: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/2.jpg)
![Page 3: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/3.jpg)
Safety concerns prior to commercialization:stability of the genetic modification, gene transfer to related plants, gene transfer to unrelated organisms, weediness potential, secondary and non-target adverse effects
3
identify + evaluate risk
release and cultivation of these plants traditional plants.
![Page 4: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/4.jpg)
4
![Page 5: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/5.jpg)
In order to be able to assess the environmental safety of GM plant, one must be:
5
1. Familiar with the biology of the plant
2. Familiar with the novelty trait
3. Familiar with agricultural/silvicultural practices employed it its cultivation
![Page 6: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/6.jpg)
Can be used to identify species-specific characteristics that may be affected by the novelty trait
6
Provide details on significant interactions between the plant and other life forms
![Page 7: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/7.jpg)
Environmental risk of GM plants
7
Novelty trait of the genetic elements
Used to transform it
Novelty of the plant itself
![Page 8: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/8.jpg)
Changes in agricultural practices associated with the introduction of new plant varieties, including, but not limited to, transgenic plants, can have significant environmental impacts.
8
![Page 9: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/9.jpg)
Requires:sufficient analytical data be available through experiment for effective comparison
9
Goal:not to establish an absolute level of
safety but a relative level of safety
![Page 10: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/10.jpg)
10
Zea mays
![Page 11: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/11.jpg)
11
No difference in seed/plant maturity observed
No significant difference in yield
No difference in agronomic quality, disease, insect susceptibility
Other than European Corn Borer Control
![Page 12: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/12.jpg)
MON 810 contains DNA sequences from the ff:
12
1. Bacillus Thuringiensis cry1Ab gene-modification of the native gene-truncated
2. Cauliflower mosaic virus (CaMV) enhanced 35s promoter
-with duplicated enhancer region
3. intron from maize hsp70 gene (heat shock protein)
-increased level of transcription
![Page 13: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/13.jpg)
4. the 3’ untranslated region of the nopaline synthase gene
-from Ti plasmid of agribacterium tumefaciens-provides polyadenylation signal for stable
expression
13
None of them are known to have any Pathogenic characteristics
![Page 14: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/14.jpg)
The following are present only in plasmids for transformation process:
14
1. CP4EPSPS marker gene2. Glyphosate oxidase marker gene3. Neomycin Phospotransferase II encoding
bacterial marker gene4. Lac Z5. ori-pUC6. CTP 2
![Page 15: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/15.jpg)
Potential Pathogenicity of the Donor Organism
15
-produce crystalline insecticidal proteins (delta endotoxins)
-humans are not affected-no deleterious effects to non-target
organisms
HOW DOES Cry1Ab WORK?
![Page 16: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/16.jpg)
(insect gut)Protein binds to specific
receptors
Inserts into membrane
Forms cation-specific pores
DEATH and PARALYSIS
16
![Page 17: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/17.jpg)
17
Why is there a need to identify the potential pathogenicity of the donor organisms? (relation to ERA)
- To identify if it has significant effects to humans, non-target animals
![Page 18: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/18.jpg)
2. Microprojectile Transformation-for direct delivery of DNA to host genome- had intact copies of the transgene, as well
as multiple, rearranged,and/or truncated transgene fragments
HOW it WORKS?
18
Plasmid w/ gene of interest
Tungsten/gold particles
(microcarriers)
Host cell
(nucleus)DNA may separate with
microcarrierIntegrate with host
genome
1. Agrobacterium mediated transformation
![Page 19: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/19.jpg)
Description of the transformation method (MON 810)
-microprojectile bombardment -although DNA solution contained genes that
code for glyphosate tolerance (CP4 EPSPS), no such gene in MON 810 genome* glyphosate selection
19
![Page 20: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/20.jpg)
While information on the integrity and copy number of the inserted DNA are generally required by regulatory authorities, there is no evidence to suggest that transgenic plants containing multiple copies of the inserted DNA are any less safe than comparable plants containing only a single copy.
![Page 21: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/21.jpg)
-number of copies of the inserted DNA*MON 810 contains 1 fragment of integrated DNA
-compositon and integrity of the inserted DNA* MON810-produced by PAT with DNA solution composed of cry1Ab, CP4 EPSPS, gox and nptII genes
21
MON 810:
![Page 22: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/22.jpg)
Why is there a need to conduct a Molecular Characterization of the inserted DNA?
-to convey that plants with more than 1 transgene copy aren’t less safe than those with only 1.
-to address issues related to positional effects, pleiotropic effects, and gene silencing
![Page 23: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/23.jpg)
Segregation Analysis of MON 810 -segregation and stability data are consistent
with a single active site of insertion of the cry1Ab gene into genomic DNA of maize line MON 810
Stability of insertion: demonstrated through 7 generations of crossing Integron analysis
23
Integron Analysis of MON 810 -Southern blot analysis: cry1ab insertion
stable in 3 generations of MON 810 breeding
![Page 24: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/24.jpg)
![Page 25: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/25.jpg)
TRIALS
ANTISERA
![Page 26: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/26.jpg)
Field Trials
US and European field trials assessed Cry1Ab, CP4 EPSPS, and GOX protein levels in maize tissues using validated ELISAs and Western Immunoblot Analysis specific for each protein.for both trials, ELISAs for CP4 EPSPS and GOX proteins were not performed since the genes are not present in maize fine MON 810.the Cry1Ab protein levels in MON810 are expressed in low level. The Cry1Ab protein in line MON 810 shows immunoreactive product of the expected size when compared with the purified protein standard.
![Page 27: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/27.jpg)
![Page 28: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/28.jpg)
Analytical technique used to detect specific proteins in a given sample of tissue homogenate or extract. It provides more specific results than the ELISA protocol.
There’s more..There’s more..
![Page 29: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/29.jpg)
![Page 30: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/30.jpg)
![Page 31: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/31.jpg)
![Page 32: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/32.jpg)
For gene flow to occur via normal sexual transmission, certain conditions must exist:
![Page 33: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/33.jpg)
![Page 34: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/34.jpg)
Why is there a need to know the Gene transfer to Related plants?
- To assess environmental risks associated with outcrossing from transgenic plantsknow reproductive biology of plant, distribution of sexually compatible relativesimpact of introduced trait.
Up next: Gene Transfer to
unrelated Plants organisms
![Page 35: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/35.jpg)
Gene Transfer to Unrelated Organisms
Horizontal gene transfer (HGT)
non-sexual exchange of genetic material between organisms belonging to the same, or different species.
HGT between plants & bacteria has been seen as a risk associated with genetically engineered (GE) plants (possible transfer of gene encoding antibiotic resistance).
![Page 36: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/36.jpg)
Up next: weediness potential
Consequences & Environmental Impact of HGT
•The transference of antibiotic resistance traits to bacteria, in either the soil or gut from transgenic plants has often been cited as a potential adverse outcome of HGT.
BUT….•-MON810 has no novel phenotypic characteristics that would extend its use beyond the current geographic range of maize production.
THUS…
•-risk of transferring genetic traits from MON810 maize to species in unmanaged environments was insignificant.
![Page 37: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/37.jpg)
![Page 38: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/38.jpg)
Weeds = subset of plants = considered pests
used to describe a plant that is a nuisance in managed ecosystems such as farms or forest plantations.Typically weeds are plant species that spread easily in disturbed areas or among crops
THEREFORE…………………
THEREFORE…………………
![Page 39: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/39.jpg)
Weediness potential is really a measure of a plants ability to successfully colonize an ecosystem, especially when it may also lead to the displacement of other species.
In the case of MON810 maize…. No competitive advantage was conferred to it
other than resistance to ECB. Does not render maize weedy or invasive of
natural habitats since none of reproductive or growth characteristics were modified.
Unlikely to establish in non-cropped habitats No reports of maize surviving as a weed
Thus.. Zea mays is not invasive and is a weak
competitor with very limited seed dispersal.
Up next: secondary and non-target adverse
effects
![Page 40: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/40.jpg)
Secondary and Non-Target Adverse Effects
Environmental risk assessment must consider the unintended consequences of the environmental release of a transgenic plant, particularly as this may impact on existing agricultural and the agro-ecosystem.
In case of plant pesticides, EPA’s intent is to evaluate potential hazard to……. ~terrestrial wildlife ~aquatic animals ~plants and ~beneficial insect
![Page 41: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/41.jpg)
Non-Target Test Organisms*Avian test species
*Aquatic animals … A. Freshwater fish species B. Aquatic invertebrate
species*Non-target insect testing
Choice of appropriate indicator organisms is based on potential for….
#field exposure to novel protein expressed in transgenic plants
#EPA has considerable background data #standard method for care and handling are
available #species are widely distributed (available) #have variety of food habits and habitat
requirements
![Page 42: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/42.jpg)
Plant pest potential
A plant may be considered a pest but not a weed
For example, a plant that produces an allelopathic substance may be considered a pest if the toxin produced has an undesirable environmental effect. Transgenic plants expressing novel toxins or potential allergens must be assessed accordingly.
![Page 43: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/43.jpg)
Effects on Non-target organisms
Bacillus thuringiensis subsp. kurstaki (B.t.k.) contains the Cry1Ab protein used to produce MON 810
History of use bacterial BT protien is not toxic
to humans, other vertebrates and beneficial insects.
only active against specific lepidopteran insects
Field Studies: United States
![Page 44: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/44.jpg)
Field trials were conducted in the United States from 1993-95 to assess the impact of insect protected maize on beneficial arthropods. Maize inbreds and hybrids expressing the Cry1Ab protein were compared to their non-transformed counterparts for relative abundance of beneficial arthropods.
Studies demonstrated that CRY1Ab had neither a direct nor indirect effect on the beneficial arthropod populations. Field trials conducted at France also have the same result. #
45.
Slide 45
![Page 45: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/45.jpg)
Laboratory Studies
-Honey bee larvae & adults -Collembola
-Parasitic Hymenopteran -Daphnia-Ladybird beetles -Green
lacewing -Northern Bobwhite Quail -Earthworm
In all cases, there were no observavble adverse effects.
MON810 maize did not present an increased risk to or impact on interacting organisms, including humans with the exception of specific lepidopteran insect species.Up next: Insect
Resistance Management
![Page 46: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/46.jpg)
Insect Resistance Management
Bt toxin control a range of insect pests including the European corn borer (ECB), cotton bollworm and the Colorado potato beetle.
![Page 47: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/47.jpg)
Bt microbial insecticides have an enviable history of safe use thus, regulatory authorities in the US and Canada consider Bt toxins in microbial sprays and transgenic plants to be in the public good and therefore worthy of extra regulatory protection.
In order to prolong the effectiveness of plant-expressed BT toxins and the microbial spray formulations of these same toxins…..
+INSECT RESISTANCE MANAGEMENT+ programs
![Page 48: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/48.jpg)
Insect Resistance Management Plans
high dose/refuge strategy means of delaying the onset of Bt resistance. This strategy involves exposing a portion of the pest population to Bt plants with an extremely high concentration of toxin [25 times the amount needed to kill 99% of the susceptible insects while maintaining another part of the population in a refuge where the pests do not encounter any Bt toxin.
![Page 49: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/49.jpg)
Fig. 1. How the high-dose/refuge strategy works to delay the increase in highly resistant (RR) insects in a pest population. Reproduced fromCohen et al. 2000.
BT CROP NON-BT CROP
4 essential assumptions:
![Page 50: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/50.jpg)
1. Resistance genes must be nearly recessive.
2. The genes conferring resistance are rare.
3. The refuge of non-transgenic plants will maintain a sufficient number of susceptible
individuals (SS homozygotes) to outnumber the resistant individuals (RR homozygotes) during mating so that their offspring will be RS heterozygotes, thus maintaining a population that is almost exclusively RS or SS.
4. The proximity of refuges to Bt crops is sufficient to ensure nearly random mating within the typical dispersal distance of the adults.
![Page 51: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/51.jpg)
Why need IRM plan?
A specific IRM plan is necessary to ensure long term resistance management
IRM strategies should be tailored to address specific regional resistance management concerns, as appropriate.
Essential for sustainable pest management goals
![Page 52: Final report mst 5](https://reader036.fdocuments.us/reader036/viewer/2022062418/555bb84fd8b42a0d0f8b4db7/html5/thumbnails/52.jpg)
Tapos na po.Salamat.