What Temperatures Are Involved in the PCR Cycle and What Processes Take Place at Each Temperature?
The polymerase chain reaction (PCR) cycle consists of three main steps, each occurring at a specific temperature:
1. Denaturation (95°C) - DNA strands are separated by breaking the hydrogen bonds between complementary base pairs.
2. Annealing (55-70°C) - Primers bind to complementary sequences on the single-stranded DNA, allowing DNA polymerase to attach.
3. Extension (72°C) - DNA polymerase extends the primer by adding complementary nucleotides, synthesizing a new DNA strand.
Additional temperatures may be included in the PCR cycle, such as:
- Initial denaturation (98°C): Activates heat-resistant DNA polymerase enzymes.
- Final extension (72°C): Ensures complete extension of all DNA fragments.
Related Questions and Answers:
- What is the purpose of the denaturation step? To separate DNA strands.
- What determines the annealing temperature? The melting temperature of the primers.
- What enzyme carries out extension? DNA polymerase.
- What is the optimal extension temperature? Typically 72°C.
- What is the purpose of the initial denaturation step? To activate DNA polymerase enzymes.
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