Question

What Reactions Are Involved In Copying Genes In PCR?

Answer 1

1. PCR or Polymerase Chain Reaction is a technique used in molecular biology to create several copies of a certain DNA segment.
2. This technique was developed in 1983 by Kary Mullis, an American biochemist.
3. PCR has made it possible to generate millions of copies of a small segment of DNA this tool is commonly used in the molecular biology and biotechnology labs.

The PCR involves three major cyclic reactions:

Denaturation:

1. Denaturation occurs when the reaction mixture is heated to 94℃ for about 0.5 to 2 minutes this breaks the hydrogen bonds between the two strands of DNA and converts it into a single-stranded DNA.
2. The single strands now act as a template for the production of new strands of DNA the temperature should be provided for a longer time to ensure the separation of the two strands.

Annealing:

1. The reaction temperature is lowered to 54-60℃ for around 20-40 seconds. Here, the primers bind to their complementary sequences on the template DNA.
2. Primers are single-strand sequences of DNA or RNA around 20 to 30 bases in length.
3. They serve as the starting point for the synthesis of DNA.
4. The two separated strands run in the opposite direction and consequently there are two primers- a forward primer and a reverse primer.

Elongation:

1. At this step, the temperature is raised to 72-80℃ the bases are added to the 3’ end of the primer by the Taq polymerase enzyme.
2. This elongates the DNA in the 5’ to 3’ direction the DNA polymerase adds about 1000bp/minute under optimum conditions.
3. Taq Polymerase can tolerate very high temperatures. It attaches to the primer and adds DNA bases to the single strand as a result, a double-stranded DNA molecule is obtained.