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can you explain the stepwise process of PCR?

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Step 1: Denaturation

As in DNA replication, the two strands in the DNA double helix need to be separated. The separation happens by raising the temperature of the mixture, causing the hydrogen bonds between the complementary DNA strands to break. This process is called denaturation.

Step 2: Annealing

Primers bind to the target DNA sequences and initiate polymerisation. This can only occur once the temperature of the solution has been lowered. One primer binds to each strand.

Step 3: Extension

New strands of DNA are made using the original strands as templates. A DNA polymerase enzyme joins free DNA nucleotides together. This enzyme is often Taq polymerase, an enzyme originally isolated from a thermophilic bacteria called Thermus aquaticus. The order in which the free nucleotides are added is determined by the sequence of nucleotides in the original (template) DNA strand. The result of one cycle of PCR is two double-stranded sequences of target DNA, each containing one newly made strand and one original strand. The cycle is repeated many times (usually 20–30) as most processes using PCR need large quantities of DNA. It only takes 2–3 hours to get a billion or so copies.

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Bishnu Parida

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