1. Genetic Engineering and Biotechnology
4.4

2. 4.4.1 Polymerase Chain Reaction (PCR)
·PCR is the cloning of DNA (amplification)
·Amount of DNA can be increased rapidly
·Useful if the source of DNA is small
·Temperature is used instead of enzymes (helicase) to separate the DNA strands
·DNA polymerase is thermostable (not denatured by the heat) and is still able to add new nucleotides to the parent strands
·This is an automated process that requires specialized machines called ‘Thermocyclers”

3. Animation:
Click techniques, then amplifying, then PCR animation

4. 4.4.2
Gel Electrophoresis
·Agar gel with rectangular indentations called “wells” at one end between a positive and negative electrode.
·DNA that has been cut into fragments (by an enzyme) is mixed into a solution with dye and then placed into one of the wells
·An electrical current is passed across the gel.

5. 4.4.2
·DNA fragments are separated based on their size (how many base pairs long they are) and charge
·Since DNA is a negatively charged molecule all of the fragments will move towards the positive electrode (which is why the wells are always located nearest the negative electrode)
- Smaller fragments are the easier to move through the gel and therefore will move more quickly to the right.
- Large fragments move more slowly.

6. 4.4.2 ·The picture below is of a gel after the DNA has been separated
·Shows 6 separate samples of DNA
·Each band corresponds to a group of DNA molecules of the same size
·Samples b and c are identical and must therefore be from the same person

7. 4.4.3 Gel electrophoresis is used for DNA profiling
·Satellite (Tandem repeating) DNA are highly repetitive sequences of DNA from the non coding region of DNA.
·Different individuals have a unique length to their satellite regions.
·These can be used to differentiate between one individual and another.
·There are different types of 'DNA fingerprinting' for different circumstance
- Forensic crime investigations
- Parentage Issues
- Animal breeding pedigrees
- Disease detection b

8.
·Bands from different samples of DNA are compared to see if they match up
·In the case of forensic investigations, one sample would be from the crime scene, one from the victim, and one from the suspect
·If the bands match perfectly the suspect can be linked to the crime scene
·In the case to the right suspect #1 appears to have been at the crime scene (must still be proven by law to have committed the crime)

9. 4.4.5
·In the case of paternity suits a sample of the child’s DNA, the mother’s DNA, and any potential father’s DNA is compared
·About half of the bands in the child’s DNA sample will match with the mother
·If the other half match with that of the third sample, the male is the father
·In the example to the right Male #1 is the father
paternity animation

10. 4.4.6
Human Genome Project
***NOTE*** this information was provided in a handout
·Government funded project that began formally in 1990 and was completed in 2001
·Purpose was to:
- Identify all the approx 30,000 genes in the human genome
- Determine the sequences of the 3 billion base pairs that make up the human DNA
- Store this information in a database

11. 4.4.6 ***NOTE*** this information was provided in a handout
- Improve tools for data analysis
- Transfer related technologies to the private sector
- Address the legal, ethical, and social issues (ELSI) that would arise from the project
* e.g. Would people really want to know if they carried the gene for a lethal disease?
* e.g. Should humans be cloned?
·To help achieve these goals, researches also studied the genomes of other organisms such as e. coli bacteria, the fruit fly, and the lab mouse

12. ***NOTE*** this information was provided in a handout

13.
Gene transfer
·The genetic code is universal (all organisms on earth use the same nucleotides)
·Genes can therefore be easily transferred between different organisms
·Genes from one organism that have been inserted into the DNA of another will still be transcribed and then translated into the correct protein

14. 4.4.8 ***NOTE*** this information was provided in a handout
Process involves:
·Restriction enzymes cut DNA at specific sites on each end of the desired gene
·At each end of the cut out gene are ‘sticky ends’ where there are hydrogens that will easily bond

15. 4.4.8 ***NOTE*** this information was provided in a handout
·Gene is inserted into a plasmid of an e. coli bacterium by cutting it open at certain sites (called restriction sites) and allowing the ‘sticky ends’ of the gene to bond
·The enzyme ligase will help to seal in the new gene
·Bacterium with plasmid can now produce the protein that the newly inserted gene codes for
·Bacterium will replicate itself numerous times (through binary fission) until there is a colony of bacterium that express the new gene
·One example of how this is used is the creation of human insulin for diabetics

16. 4.4.8
***NOTE*** this information was provided in a handout

17. 4.4.9 Genetically Modified Organisms (GMO’s)
·GMO is an organism carrying a transplanted gene
·The organism will express the gene and synthesize proteins from it
·One example is the production of human clotting factor by genetically modified sheep
- The clotting factor is isolated from the sheep’s milk and then used to treat hemophiliacs

18. 4.4.9
·Other examples include pest resistant crops, salt tolerant tomatoes, and rice that produces beta carotene (a precursor to vitamin A)

19.
Click on link "Should we grow GM crops?"
Take the survey
Outline 3 pro and 3 con arguments.
Explain your position.
Due Monday 2/14/11

20. 4.4.10 ·The creation and use of GMO’s is a highly debated topic:
·Benefits
·Increased crop yields, particularly in regions of food shortage
·Yield of crops with specific dietary requirements such as vitamins and minerals (esp. good for malnourished and poverty stricken regions of the world)
·Crops that do not spoil easily during storage or transport
·GM animals may provide higher meat yields

21. 4.4.10
·Disadvantages
·The foods (animal and plant) are considered unnatural and unsafe for human consumption by some people
·There is the risk of the escape of genes into the environment where they may be passed to other organisms with unknown side effects (e.g. passing pest resistance on to a fast growing weed)

22.
Cloning
·Clone = a group of genetically identical organisms or a group of cells derived from a single parent cell
·The first successful cloning of a mammal was Dolly the sheep. The process for this cloning is as follows:
·Isolate the donor nucleus from a somatic cell of a donor sheep
·Retrieve unfertilized eggs from a female sheep. Many eggs are needed since not all will survive the process

23. 4.4.12
·Remove the egg cell’s nucleus, which contains only half of the sheep’s genetic material (a very small needle and syringe is used to poke through the cell membrane)
·Insert the nucleus, complete with the DNA that it contains, into the donor cell that has no nucleus
·Place the egg into a female sheep’s womb. Only a small percentage of the eggs placed in the womb will continue to develop into embryos
·When the offspring is born it will be a genetic clone of the sheep that donated it’s nucleus

24. 4.4.12

25. 4.4.13 ·Cloning humans presents many ethical issues ·Proponents say:
·Cloning happens naturally, for example monozygotic twins (identical twins).
·Cloned embryos can be tested for genetic abnormalities
·Cloned embryos increase the chances of having a child for infertile couples

26. 4.4.13
·Opponents say:
·Costly process; money better spent on other health procedures
·Cloned individuals are more likely to have developmental problems
·May be done for innapropriate reasons (replace a loved one, build a “super race”, etc)