1. Transgenic Plants Production of Bovine Somatotrophin
Agriculture
Transgenic Plants
Production of Bovine Somatotrophin

2. Revision
How is genetic engineering of organisms carried out (think back to SG and Higher)?
What is needed?
What is substances are produced through genetic engineering?
What else do we know?

3. Transgenic Plants
A transgenic organism is an organism that is carrying genetic material of another organism in its genome.
Producing a transgenic organism requires:
A vector used to insert the required DNA into a plant
A way of ensuring the gene is carried in all cells
Discuss here what knowledge we already have of this from plant cell tissue culture.

4. Vectors
A vector is an agent used to transfer DNA from one organism to another
Agrobacterium (tumerfaciens) is a commonly used vector
Agrobacterium is a soil bacteria
It contains a plasmid (known as Ti) which it can insert into plant DNA causing a tumour (crown gall disease)

5. Vectors cont…
Compare to a virus – the DNA becomes integrated into the plant DNA, but it doesn’t take over the cell and destroy it. Instead it causes the production of plant hormones and proteins that act as nutrients for the bacteria. As a result there is positive feedback – the cells grow more, with the bacterial DNA being copied at each division and more bacterial growth. The uncontrolled growth results in a lumpy cell mass – the gall. At the time they were working out the mechanism of Agrobacterium (1970s) it was noted that if a human was to do this artificially a P4 containment facility would have been required. In other words the agrobacterium was working outside the national institutes of Health guidelines.

6. Procedure for creating a transgenic plant using a Ti-plasmid
Ti disease gene is disabled
Eg. By removing it with restriction enzymes
A target gene containing desired characteristics is identified
A target gene is removed from source DNA using a restriction enzyme
The plasmid is then cut using the same restriction enzyme

7. Procedure for creating a transgenic plant using Ti cont…
The foreign DNA (target gene) is then added to the plasmid and sealed using DNA ligase
The modified plasmid is returned to the Agrobacterium
(Note the plasmid also has a method for identification e.g antibiotic resistance – later in the process this is used to identify the cells that have the plasmid)

8. Procedure for creating a transgenic plant using Ti cont…
Plant cell protoplasts are prepared
The protoplasts are incubated with the bacterium containing the modified plasmid
Grown in a selective medium that allows only the growth of cells containing the plasmid (and selected gene)
E.g the medium may contain an antibiotic

9. Example 1 – insect resistance
The bacteria Bacillus thuringiensis produces proteins (known as BT toxins) that have been used for insecticides
The toxin is isolated from the bacteria genome and inserted into Ti plasmid of agrobacterium
The resulting plant kills insects that eat it
Discuss the issue of most pesticides being harmful to all animals including the targeted insects. BT was first identified in 1901 when it devastated Japan’s silkworm population. It damages the gut of insects, leaving the gut paralysed and insects die from tissue damage and starvation. Benefit is the bacteria is destroyed in low pHs of most animal guts, but thrives in the alkaline conditions found in insect guts.

10. Example 1 – insect resistance
Potato, cotton and soybean are all plants were this technique is used
Benefits
Only insects eating the plant are killed
Reduces the use of pesticides
The entire plant has protection (not just the leaves and stem, as would be the case if it was sprayed)

11. Example 2 – Tomato plants
Tomato ripening is naturally accompanied by softening due to an enzyme – PG (polygalacturonase)
Fruit an usually picked while still green and ripened artificially using ethene gas
By modifying the PG gene, fruit could be left to ripen longer without softening
Fresher, riper tomatoes could be delivered to supermarkets
PG does this by breaking down the cell walls
Used to produce flav-savr – a commercial disaster due to the taste – a poorly flavoured tomato was used in the first place

12. Bovine Somatotrophin (BST) production
Bovine somatotrophin is a growth hormone that can be used to increase growth (mainly muscle and bone) and milk production in cattle
The gene is isolated from cattle cells using restriction enzymes
A restriction enzyme is also used to open a plasmid from an E. coli bacteria

13. Bovine Somatotrophin (BST) production
The BST gene is inserted into the plasmid using DNA ligase
The E.coli bacteria containing the plasmid are then cultured
The gene is expressed (i.e. transcribed and translated into protein form) during bacterial growth
The protein is then purified and prepared for administration to cattle
Discuss the use of antibiotics again. DART goes into explicit detail about the use of Southern blotting and probes to identify – too much info me thinks

14. Use of BST
BST is either administered by injection or the protein is included in cattle feed
Results in a 10% increase in milk production
BST results in the stimulation of protein synthesis and cell division because it encourages the uptake of glucose and amino acids

15. Disadvantages / concerns
Can increase mastitis in cows
Lameness later in life – due to enlarged udders
BST causes a lack of fat deposits (more of the glucose etc.. is used) therefore hypothermia can develop
BST found in milk – supposedly safe as it will be broken down in our stomachs
Cows are already producing greater milk quantities than they used to, so the addition of BST can highlight this

16. Learning Activities Monograph pg 90 – 94 DART pg 88 – 93 Scholar
Genetically Modified Plants worksheet
Make flow diagrams / posters of the process of
Creating transgenic plants
BST production
Assignment Questions – 2001 Q12, 2005 Section B Q 8
Look at websites

17. Moral and Ethical Issues related to transgenic organisms
Moral and Ethical codes attempt to establish acceptable human behaviour for the benefit of all society ‘DART’
Transgenic organisms
What rights do they have?
Farmers
What might there opinions towards transgenic be? How might differing views on accompanying farms be dealt with?

18. Moral and Ethical Issues related to transgenic organisms
Consumers
What are the pros and cons?
What are the facts vs media hype?
The Environment
Benefits vs problems
Biotechnology Industry
Motives vs issues

19. Formal Debate
This house supports the creation and use of transgenic organisms

20. THE END OF UNIT ONE
(Except for the NAB)