Question

The que Carries {4 marks}.

Q] Explain in detail polymerase chain reaction?

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Answer 1

Answer:

PCR involved a process of heating and cooling called thermal cycling which is carried out by machine.

Three steps occur:

1) DENATURING : when the double standard template DNA is heated to seperate it into single strands.

they occur 94-95°C

2) ANNEALING: when the temperature is lowered to enable the DNA primers to attach to the template DNA.

they occur 50-56°C

3) EXTENDING: when the temperature is raised and the new strand of DNA is made by the taq polymerase enzyme.

These three stages are repeated 20-40 time, doubling the number of DNA copies each time.

they occur 72°C

Answer 2

Polymerase Chain Reaction :

In many situations, there is only a small amount of DNA available for DNA fingerprinting. If needed many copies of the DNA can be produced by PCR (DNA amplification).

Process Of PCR :

There are three steps for amplification of a gene sample of interest carried out using PCR. They are :

1. Denaturation
2. renaturation or primer annealing
3. synthesis or primer extension.

• The double stranded DNA of interest is denatured to separate into two individual strands by high temperature . This is called denaturation.
• Each strand is allowed to hybridize with a primer (renaturation or primerannealing). The primer template is used to synthesize DNA by using Taq – DNA polymerase.
• During denaturation the reaction mixture is heated to 95 °C for a short time to denature the target DNA into single strands that will act as a template for DNA synthesis.
• Annealing is done by rapid cooling of the mixture, allowing the primers to bind to the sequences on each of the two strands flanking the target DNA.
• During primer extension or synthesis the temperature of the mixture is increased to 75°C for a sufficient period of time to allow Taq DNA polymerase extend each primer by copying the single stranded template.
• At the end of incubation both single template strands will be made partially double stranded. The new strand of each double stranded DNA extends to a variable distance downstream.
• These steps are repeated again and again to generate multiple forms of the desired DNA. This process is also called DNA amplification.