“The polymerase chain reaction (PCR) is a powerful technique used in biology. Principally, PCR is based on amplification of DNA fragments of interest using a specific thermostable DNA polymerase (Taq polymerase) that replicates the target DNA sequence. At the end of the process, the target sequence is multiplied several million times. Thanks to this fact, the technique enables amplification of trace amounts of the sample.

Besides Taq polymerase and target DNA, the reaction requires a pair of primers, dNTPs, and monovalent and divalent cations (e.g. Mg²⁺). The primer is a short oligonucleotide, usually 18-24 base pairs in length. A pair of primers is designed in such a way as to match to the beginning and the end of DNA the fragment of interest. After the primers anneal to the fragment, Taq polymerase can add new nucleotides at the 3′ end of the DNA fragments. PCR is performed on thermocyclers which automate specific temperature cycles. A standard PCR procedure consists of 25 to 40 cycles, where each cycle consists of the following three steps:

1. Denaturation. This step involves heating of the reaction to 94-98 °C for 30-45 seconds. This leads to unwinding of the double-stranded DNA molecule and its separation into two single-stranded molecules.
2. Annealing. During this step, the reaction temperature is lowered to 50-65 °C for 30-60 seconds. This allows primers to anneal to the target DNA sequence. Stable DNA-DNA hydrogen bonds are only formed when the primer sequence very closely matches the template sequence
3. Extension. The temperature is increased to 72 °C and Taq polymerase begins to synthesize a new DNA strand complementary to the DNA template strand by adding dNTPs to the primer’s 3′ end.

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