AAT Bioquest

What are the 4 steps of PCR?

Posted May 18, 2020


PCR consists of the four following steps:

  1. Denaturation by Heat: double-stranded DNA is separated into two single strands, by a process called denaturation which occurs at temperatures higher than 90 degrees Celsius. Heat breaks hydrogen bonds between the base pairs, while the stronger bonds between deoxyribose and phosphates, remain intact.
  2. Annealing Primer to Target Sequence: before the target sequence (generally between 100-600 base pairs) is replicated, it must be targeted using primers that target the ends of the target DNA sequence by annealing (binding) to the complementary sequence. Annealing occurs at lower temperature (between 40 and 65 degrees Celsius) which depends on the length and base sequence of the primers.
  3. Extension: after annealing, the temperature is increased to 72 degrees Celsius and the Taq DNA polymerase enzyme is used for replicating the DNA strands. During synthesis (or extension), two identical double stranded DNA molecules are synthesized.
  4. End of the First PCR Cycle: after which the process is repeated generally between 25 and 35 times.