Question

Describe the steps of PCR technique.

Answer 1

Hint: PCR is also referred to as polymerase chain reaction. By using PCR technique, amplification of a DNA segment can be done easily by following a few steps. For performing PCR, ingredients such as Taq polymerase, primers, template DNA and nucleotides are required. These are mixed together in a PCR tube.

Complete answer:
There are 3 main steps involved in PCR
> Step 1: Denaturation- The two DNA strands are to be separated and hence this process is necessary. The temperature of the mixture is raised which causes hydrogen bonds present between complementary DNA to break. 94 degree Celsius heat has to be applied in this step.

> Step 2: Annealing- Binding of the target DNA sequence to the primers will happen. This starts the polymerization process. Here, the temperature has to be lowered to 50-60 degree Celsius for the primer to bind to each strand.

> Step 3: Extension- This is the last step. New DNA strands can be made using original ones. DNA polymerase enzyme joins DNA nucleotides together. Taq polymerase is most widely used. It is isolated from a bacteria called thermus aquaticus. The free nucleotides are added in a specific order of original template DNA strand.

Note: The template DNA is isolated from organisms containing a target region. DNA primers are short pieces of single stranded DNA. PCR has many important applications such as paternity testing, genotyping, to detect a mutation, in genome sequencing and many others. There are many PCR mistakes that happen which lead to false results. Some of which are; primer design is not appropriate, cross contaminations or the reaction efficiency being more. There is a huge list of PCR types. Some of which are colony PCR, allele-specific PCR, conventional PCR, hot start PCR and multiplex PCR.