PCR is based on ability of DNA polymerase to synthesize complementary strand to the template strand. As DNA polymerase can add a nucleotide only onto a 3'-OH group, it needs an artificial DNA strand (called DNA primer) of about 18 to 25 nucleotides complementary to 3' end of the DNA template. As shown below, each polynucleotide has a free 3' −OH group and 5' phosphate group. Moreover, a DNA strand has complimentary sequence, already paired by hydrogen bonding. Thus, primer can bind only when DNA strands are separated. This is generally done by heating.

The primers anneal to the single-stranded DNA template at specific temperature (depends on primer sequence) and then DNA-Polymerase binds to this double stranded DNA produced. Then again reaction mixture is heated to 72°C (e xtension ); a temperature optimum for DNA- polymerase functions. This starts synthesis of the new DNA strand. Then reaction mixture is cooled to lower temperature for short term storage, if required. This completes one cycle. After first cycle, one DNA molecule has become two. After multiple cycle of the PCR reaction, the specific sequence will be accumulated in billions of copies.

The PCR reaction requires the following components:

DNA template : DNA template is DNA target sequence. As explained earlier, at the beginning of the reaction, high temperature is applied to separate both the DNA strands from each other so that primers can bind during annealing.

DNA polymerase: DNA polymerase sequentially adds nucleotides complimentary to template strand at 3'-OH of the bound primers and synthesizes new strands of DNA complementary to the target sequence. The most commonly used DNA polymerase is Taq DNA polymerase (from Thermus aquaticus, a thermophillic bacterium ) because of high temperature stability. Pfu DNA polymerase (from Pyrococcus furiosus ) is also used widely because of its higher fidelity (accuracy of adding complimentary nucleotide).
Mg²⁺ ions in the buffer act as co-factor for DNA polymerase enzyme and hence are required for the reaction.

Primers: Primers are synthetic DNA strands of about 18 to 25 nucleotides complementary to 3' end of the template strand. DNA polymerase starts synthesizing new DNA from the 3' end of the primer.