Genetic Modification and Biotechnology

Polymerase chain reaction (PCR). Method to make millions of copies of a specific DNA region. Each cycle (~2 minutes) doubles the amount.

Steps per cycle:

1. Denaturation (~95 °C) — heat separates DNA strands.
2. Annealing (~55 °C) — short DNA primers bind to complementary sequences on each strand.
3. Extension (~72 °C) — Taq polymerase (heat-resistant DNA polymerase from Thermus aquaticus) extends primers, synthesising new strands.

After 30 cycles: 2³⁰ ≈ 10⁹ copies of the target sequence.

Applications: forensic DNA profiling, prenatal genetic testing, diagnosis (e.g. COVID-19 RT-PCR), ancestry analysis, ancient DNA studies.

Gel electrophoresis. Separates DNA fragments by size.

1. Sample DNA cut with restriction enzymes → fragments of various sizes.
2. Loaded into wells in an agarose gel; current applied.
3. DNA is negatively charged (phosphate groups) → migrates toward positive electrode.
4. Smaller fragments move FURTHER (less resistance through gel).
5. Stained (e.g. with fluorescent dye) and visualised under UV.

DNA profiling. Pattern of fragments produced by cutting tandem repeat regions (short tandem repeats — STRs) varies between individuals. Used in paternity testing and forensics — match probability commonly < 1 in a billion when many STR loci are compared.

Recombinant DNA technology.

1. Identify the gene of interest (e.g. human insulin gene).
2. Cut both the gene and a plasmid (small circular bacterial DNA) using the SAME restriction enzyme → leaves matching sticky ends.
3. Mix gene + plasmid with DNA ligase → recombinant plasmid.
4. Insert plasmid into a host bacterium (transformation).
5. Host expresses the foreign gene, making the protein.

Example. Human insulin gene is inserted into E. coli. The bacteria mass-produce insulin in fermenters — cheaper and safer than the historical pig pancreas method.

Genetically modified organisms (GMOs).

Example	Modification	Benefit	Concern
Insulin-producing bacteria	Human insulin gene	Treats diabetes	None major
Golden rice	β-carotene biosynthesis genes	Reduces vitamin A deficiency	Ecological, regulatory
Bt corn	Insecticidal toxin gene from Bacillus thuringiensis	Pest resistance, less pesticide	Non-target insects, resistance

Cloning.

Reproductive cloning (e.g. Dolly the sheep, 1996):

1. Take a body cell from donor (e.g. mammary gland).
2. Take an egg cell from another sheep; remove its nucleus (enucleate).
3. Insert donor nucleus into enucleated egg → reconstructed embryo.
4. Stimulate division (electric shock); implant in surrogate.
5. Offspring is genetically identical to nuclear donor.

Therapeutic cloning — same nuclear transfer, but the embryo is not implanted. Stem cells extracted for tissue regeneration (no transplant rejection).

Natural clones. Identical twins (form from one zygote splitting); plants reproduce asexually by runners or cuttings.

Ethics.

• GMO ecology — pollen drift to wild relatives; horizontal transfer of resistance genes.
• GM patents — corporate ownership of crop genomes; farmer dependence.
• Human germline editing (e.g. CRISPR) — heritable changes affecting future generations; "designer babies".
• Animal cloning welfare — Dolly suffered premature ageing.
• Stem cells from embryos — moral status of early embryos.

These tradeoffs are tested in Paper 2 extended-response questions.