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Question
Describe three steps involved in mechanism of PCR.
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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