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Question
Describe three steps involved in mechanism of PCR.
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Solution
At the start of PCR, all the requirements are mixed together in 'eppendorf tube' and the following operations are performed sequentially:
- Denaturation: The reaction mixture is heated to a temperature (90–98° C) to separate two strands of desired DNA. This is called denaturation.
- Annealing: The mixture is allowed to cool (40–60° C) that permits pairing of the primer to the complementary sequences in DNA. This step is called annealing.
- Primer extension/Polymerization: The temperature (70–75° C) allows thermostable Taq DNA polymerase to use single-stranded DNA as template and adds nucleotides. This is called primer extension. It takes around two minutes duration.
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