Question

Steps involved in PCR (Polymerase Chain Reaction) process.

Answer 1

Steps involved in PCR process:

1. PCR process is a cycle of three successive reaction:
   1. Denaturation:

• At 93 - 95°C, the target DNA molecule is denatured, and two strands of DNA is separated.

1. Primer Annealing:

• In this step a short synthetic DNA primers are annealed to the separated strands.
• At this point the temperature must be low enough for the process of hybridisation. It must be around 50 - 70°C.

1. DNA Synthesis:

• Here polymerisation process is proceeding in presence of the enzyme DNA polymerase.
• The temperature should be about 70 - 75°C.

1. The above three steps are repeated of 25 - 30 cycles.
2. At the end of 20 cycles the number of newly synthesized DNA is 1 × 106 and after 30 cycles its 1 × 109.
3. This number can be calculated by the below formula:
   M_f = M_i × 2n
   Where,
   M_f = initial number of molecules (templates)
   M_i = final number of newly synthesized DNA molecules by PCR
   n = number of cycles took place