D1.1 PCR
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What does PCR stand for?
Polymerase Chain Reaction.
Purpose of PCR?
Amplify (copy) DNA to obtain millions/billions of copies.
Where does PCR occur?
In vitro (test tube/lab), not in a living organism.
Why is PCR powerful?
Works with tiny DNA samples (crime scenes, fossils, viruses).
Enzyme used in PCR?
Taq polymerase – heat-stable DNA polymerase.
Source of Taq polymerase?
Thermophilic bacterium Thermus aquaticus.
Why not human polymeras?
Human enzymes denature at 98°C; Taq is stable.
Three stages of PCR?
• Denaturation (98°C, strands separate). • Annealing (~60°C, primers bind). • Extension (72°C, Taq adds nucleotides).
What are free nucleotides?
Building blocks used by Taq polymerase to synthesize DNA.
What are primers?
Short single-stranded DNA that provide starting points for polymerase.
How many DNA molecules after n cycles?
2n2^n.
After 2 cycles, how many double-stranded DNAs?
4
After 25 cycles?
~33 million copies.
Real-life applications of PCR?
COVID-19 testing, forensic crime scenes, ancient DNA (Neanderthal), genetic engineering.