Lecture 11/12 - Applications of transgenic plant technology

Question 1

What are 4 examples of current applications of Transgenic technology?

Answer 1

1-herbicide tolerance e.g. introduction of tolerance to broad-spectrum herbicide Glyphosphate, ‘round-up ready’ crops
2-Pest resistance e.g. synthesis of insecticidal proteins in cotton and maize, ‘Bt crops’
3-Virus resistance e.g. using RNAi to supress viral accumulation, ‘Rainbow papaya’
4-Nutritional enhancement e.g. alteration of composition of food to increase nutritional value e.g. ‘Golden Rice’

Question 2

Why create crops with herbicide resistance?

Answer 2

-weed control one of the biggest problems in crop production and is controlled by weeding/herbicides

Question 3

What is Glyphosate?

Answer 3

a broad spectrum systemic herbicide

• does not bio-accumulate
• breaks down rapidly

Question 4

What is the process of Glyphosate treatment?

Answer 4

1) Glyphosate is added to Shikimic acid and Phosphoenol pyruvate, Plant EPSP sunthase converts to EPSP
2) forms aromatia amino acids

Question 5

How are GM Glyphosate tolerant crops ‘round up ready?

Answer 5

survive after round up treatment

-contain EBBP synthase gene from bacteria resistant to glyphosate

Question 6

What is 2,4 Dichlorophenoxyacetic acid? (2,4 D) And how is it used for herbicide resistance?

Answer 6

• synthetic auxin herbicide
• acts on auxin receptors (essential for normal plant growth)
• bacterial AAD enzyme class cleaves 2,4D
• soybeans carrying AAD transgene are resistant to 2,4 D herbicide

Question 7

What is an USA and Canadian major corn pests and what is its economical effects?

Answer 7

European corn borer (US & Canada)

-damage and control costs >$1 billion yearly

Question 8

What is Bt?

Answer 8

Bacillus thuringiensis, a soil bacterium, spores contain toxic crystalline (Cry) protein
Insecticide

Question 9

How does Bt insecticide work?

Answer 9

1) Insect eats Bt crystals and spores
2) Toxin binds to specific receptors in gut and the insect stops eating
3) gut wall breaks down allowing spores and normal bacteria to enter the body
4) Insect dies as spores and gut bacteria proliferate within the body

Question 10

Why create transgenic Bt crops?

Answer 10

Performance of Bt insecticides not consistent
-toxin is sensitive to UV, heat, dessication
-incomplete coverage of feeding sites
-reduced toxicity against older larvae
Bt crops overcome these limitations
-protein is protected from rapid environmental degredation
-plants produce protein in tissues where insects feed so coverage not an issue
-protein is present whenever newly hatched larvae emerge so timing not a problem

Question 11

What are the pros and cons of Bt crops?

Answer 11

Reduced pesticide use
-2004 farmers used 77 million kg less pesticides on Bt cotton and 6.3 million less on Bt corn than in 1993
Lowered risk of poisoning
Less danger to wildlife and environment, fewer beneficial insects harmed
Lowered costs as fewer pesticide applications may be needes
CONS
Continued exposure to insecticide can lead to insecticide resistance
Risk of losing the use of a relatively safe insecticide

Question 12

What are two examples of cotton pests?

Answer 12

Cotton bollworn

Pink Bollworm

Question 13

What is insect resistance management (IRM)?

Answer 13

Strategies to delay the development of insect resistance to Bt

Question 14

What is the most widely used IRM strategy?

Answer 14

The use of refuges

Strip of non Bt corn next to field of Bt corn

Question 15

How can refuges prolong the usefulness of Bt crops?

Answer 15

Maintains a population of insect pests not resistance to Bt proteins, allowing non resistant pests to mate with any resistance pests that might emerge

Question 16

What technique is an alternative IRM strategy to refuges?

Answer 16

Sterile Insect Technique

Question 17

What is sterile insect technique?

Answer 17

Release of sterile insects along with presence of Bt cotton to reduced the level of damage and reduce chance of moth resistance

Question 18

How has Sterile Insect technique been shown to be effective?

Answer 18

Three conditions

1) Bt cotton begins with low damage level but high resistance in absence of refuges -> emergence of resistance alleles -> severe damage from resistant insects
2) Non Bt cotton has severe damage from wild insects -> release large amount of sterile moths -> low level of damage but costly release of large numbers of sterile moths
3) Bt cotton and release of some sterile moths -> v low chance of damage with reduced change of resistance and less extensive release of sterile insects

Question 19

How is virus resistance generated in Papaya?

Answer 19

Use RNAi to supress papaya ringspot virus

Question 20

What is the push-pull system?

Answer 20

Main crop and trap crop
Main crop attracts natural enemies, moths are pushed away from the main crop
-Trap crop attracts moths

Question 21

Why is Golden Rice beneficial?

Answer 21

-worldwide, 3 billion people depend on rice as major staple
BUT rice lacks beta-carotene which can be converted to vitamin A when digested
Vitamin A deficiency major health problem
-WHO 100-140 million children are vitamin A deficient
-leading cause in preventable blindness in children
-implicated in >1million deaths yearly

Question 22

What is golden rice?

Answer 22

Rice that has been genetically engineered so that the endosperm (edible part) contain beta-carotene

Question 23

What is the B carotene pathway?

Answer 23

1) IPP converted to Geranylgeranyl diphosphate
2) Phytoene synthase converts Geranylgeranyl diphosphate to Phytoene
3) Phytoene desaturase and Carotene desaturase convert Phytoene to Lycopene
4) Lycopene-cyclase converts Lycopene to B carotene (vitamin A precursor)

Question 24

What enzymes does rice lack for making beta carotene?

Answer 24

• Phytiene synthase
• Phytodene desaturase
• Carotene desaturase
• Lycopene-cyclase

Question 25

How was golden rice created?

Answer 25

B-carotene pathway genes added

• Phytiene synthase - Daffodil gene
• Phytodene desaturase and Carotene desaturase - single bacterial gene
• Lycopene-cyclase - daffodil gene

Question 26

What is Golden Rice 2?

Answer 26

New strain of golden rice provides the daily recommended Vitamin A allowance with 70g portion rice, by using Maize Psy instead of Dafodill Psy

Question 27

Who owns golden rice?

Answer 27

• 70 intellectual propery rights (32 companies)
• Humanitarian Use license issued
• as long as farmer makes less than $10,000 no royalties paid
• farmer can save seeds for replanting

Question 28

What are 3 controversies of transgenic plants?

Answer 28

• Risk to human health e.g. antibiotic resistance genes
• Unfavourable to small farmers e.g. Terminator technology
• Risk to the environemtn e.g. monach butterfly

Question 29

How might the antibiotic resistance gene be removed from transgenic plants?

Answer 29

the Cre-lox system

Question 30

What are cre and loxP in the cre-lox system and what do they do?

Answer 30

Cre:cyclization recombination encodes a site specific recombinase
loxP: P1 bacteriophage sequence (34bp) not found in plants or animals

3) Cre catalyses recombination between loxP sites flanking selectable marker gene
1) Create transgenic plant expressing gene of interest
2) Cross plant expressing Cre recombinase
3) Cre catalyses recombination between loxP sites flanking selectable marker gene
4) Intervening DNA is excised, removing selectable marker

Question 31

How are transgenic plants unfavourable to small farmers?

Answer 31

Through use of terminator technology

Genetic Use Restriction Technology, GURT

Question 32

What is terminator technology?

Answer 32

• plant encorperates a trait (toxin) that kills developing embryos late in development
• expression of toxin is induced by a chemical applied to seed of the parent plant
• seed cannot be saved and planted next year
• prevents spread of transgenes

Question 33

How is production of the toxin prevented in seeds that have no had the chemical inducer applied? (terminator technology)

Answer 33

1) Repressor is expressed
2) repressor binds to gene binding site and prevents recombinase expression
3) blocker consequently prevents production of the toxin

Question 34

How is the toxin expressed in seeds that have had the chemical inducer applied? (terminator technology)

Answer 34

1) Repressor is expressed
2) Chemical inducer applied by seed company prevents repressor binding
3) recombinase is produced and excises the blocker
4) Toxin is available to be produced after late promotor

Question 35

Does the Bt toxin endanger pollinator species?

Answer 35

-Dusted Bt corn pollen onto milkweed leaves and fed to Monarch larvae
-Larvae ate less and grew more slowly
-over four days nearly half larvae died compared with none in the control groups
Bt 176 produces pollen that is highly toxic to Monarch larvae

Question 36

Why does Bt 176 produce a pollen that is highly toxic to Monarch larvae?

Answer 36

promotor is very efficient at producing Bt in pollen

Question 37

How has damage to Monarch butterflies been reduced from Bt corn?

Answer 37

Bt 176 (highly toxic) transgene used in <2% corn acreage
More commonly grown Bt varieties do not produce much of the toxic Bt protein in pollen
-larvae can eat 1600 grains of pollen per cm2 of leaf surface without any ill effects