Question

You are a researcher working in a molecular biology laboratory. You need to run a batch of PCRS (polymerase chain reaction) to verify the presence of a specific gene in the organism you are currently working on. 1. What are all the necessary reagents required for the PCR (hint-5)? (1 point) 2. After you assemble your reaction in the PCR tubes this goes into the thermocycler which will control the temperature necessary for each main step to occur. How are those steps called and what is their main function? (2 points) 3. After the PCR program ran to completion, you performed a gel electrophoresis. What is the function of this step? (1 points)

Answer 1

1. The necessary reagents of the PCR are template DNA, PCR primers (forward primer and reverse primer), reaction buffer, Taq polymerase and nucleotides.

2. The main steps of the PCR are:

1. Denaturation: It involves separation of the two strands of the DNA into single strands by breakage of the hydrogen bonds. It exposes the DNA nucleotides for the attachment of primers. Temperature is maintained around 95 degree celsius in the step.
2. Annealing: It includes attachment of the primers to the single-stranded DNA templates. Temperature is around 60 degree celsius in this step.
3. Extension: It involves extension of the DNA primers by addition of bases complementary to the DNA template. Temperature is around 72 degree celsius in this step. This step uses Taq polymerase.

3. After PCR, gel electrophoresis is done to visualize the results of PCR. The amplified DNA is run into gel to check their molecular weight as it separates the DNA fragments according to their size.