11.3.3 Amplification of Gene of Interest using PCR

PCR stands for Polymerase Chain Reaction. In this reaction, multiple copies of the gene (or DNA) of interest is synthesised in vitro using two sets of primers (small chemically synthesised oligonucleotides that are complementary to the regions of DNA) and the enzyme DNA polymerase.

The enzyme extends the primers using the nucleotides provided in the reaction and the genomic DNA as template. If the process of replication of DNA is repeated many times, the segment of DNA can be amplified to approximately billion times, i.e., 1 billion copies are made.

Such repeated amplification is achieved by the use of a thermostable DNA polymerase (isolated from a bacterium, Thermus aquaticus), which remain active during the high temperature induced denaturation of double stranded DNA.

The amplified fragment if desired can now be used to ligate with a vector for further cloning.

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