A-Level Biology AQA Notes

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Genetic Engineering

Genetically modified organisms are organisms that have had their DNA altered through recombinant DNA technology.
Recombinant DNA technology involves the transfer of fragments of DNA from one organism, or species, to another.
Transgenic organisms can successfully express a gene from any organism, as the genetic code and mechanism of protein production (transcription and translation) are universal.
DNA fragments are created by:
- Using restriction endonucleases to cut at recognition sites near the desired gene
- Converting the mRNA of the desired gene to cDNA, using reverse transcriptase. Double stranded DNA is then synthesised using DNA polymerase
- Synthesising the gene using a gene machine. The gene sequence is determined by the primary protein structure

The isolated gene is then modified by the addition of a promoter and a terminator region.
A vector is used to transfer the isolated gene into a host cell. This is mainly a plasmid.
Restriction endonucleases are used to cut plasmids open, creating sticky ends. The same endonuclease isolates the gene, so the sticky ends of the desired gene and the plasmid are complementary. DNA ligase joins them together
To reintroduce the desired DNA into bacterial cells, the recombinant plasmid must pass through the cell surface membrane of a bacterial cell (transformation).
Transformation involved mixing the bacteria and plasmids in a medium containing Ca2+ ions, which increased membrane permeability. Changes in temperature also make the bacterial cell surface more permeable.
The transformed host cells can be cultured as an in vivo method to amplify DNA fragments.

Marker Genes

Transformed bacteria can be detected using marker genes
The plasmid contains 2 marker genes
- The first marker gene is used to identify which bacteria have successfully taken up a plasmid. It is a antibiotic resistance gene, so transformed bacteria are identified by growing on a medium containing the antibiotic
- The second marker distinguishes between bacteria that have taken up an empty or recombinant plasmid. When a recombinant plasmid is formed, the desired gene is inserted in the middle of the second marker gene making it non-functional. Therefore, bacterial cells that express the second marker gene do not contain the recombinant plasmid

The second marker gene has easily identifiable phenotypes such as:
Producing a fluorescent protein
- Providing resistance to a different antibiotic
- Producing an enzyme whose action can be identified

Polymerase Chain Reaction (PCR)

PCR is a method of amplifying DNA by artificial replication in vitro.
It requires: DNA sample of around 10,000 base pairs, nucleotides, Taq polymerase (stable at high temperatures), primers complementary to 3’ of DNA sample and a thermocycler to carry out the automated process.

The Use of Genetically Modified Organisms (GMOs)

GMO	Benefits	Issues
Plants	Herbicide resistance Pest resistance Disease resistance Drought resistance Extended shelf-life Increased nutrition	Development of superweeds Pests or pathogens evolving resistance Potential transfer of antibiotic resistance to pathogens in the intestine of the consumer Farmers must repeatedly buy seeds
Animals	Disease resistance Increased growth rates e.g. continuously producing growth hormones Used to produce medicinal drugs and proteins	Harmful side effect to animals Ethical issue of insertion of human genes Most GM animals die during development
Bacteria	Used to produce medicine e.g. human insulin which is cheaper and has a lower risk of rejection and infection than pig insulin	Potential antibiotic resistance genes being transferred to pathogens May result in the production of more lethal pathogens

The risk of GM bacteria can be reduced by modifying the bacteria so that they are unable to produce an essential nutrient or amino acid and cannot survive outside the lab.

Gene Therapy

Gene therapy is the mechanism by which genetic diseases are treated or cured by masking the effect of a faulty allele through the insertion of a functional allele.
Firstly, a healthy allele from healthy cell tissue is isolated. The allele is inserted into the cells using vectors.
If the mutated allele is recessive, a dominant allele is inserted. If the mutated allele is dominant, DNA is inserted into the middle of the allele to silence it.
Somatic therapy involves altering the alleles in body cells. The altered allele is not passed onto the offspring
Germ-line therapy altering the alleles in the sex cells. The altered alleles are passed onto offspring
Germ-line therapy has ethical concerns such as the potential of designer babies or the potential impact gene insertion could have on other genes.

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