Polymerase Chain Reaction

Principle

Polymerase Chain Reaction (PCR) is a molecular biology method used to amplify a small amount of genetic material (DNA or RNA), using specific primers of a target sequence.
PCR is divided into 5 steps:
- First denaturation:
- Denaturation step
- Annealing step
- Elongation step
- Final step
The 2nd, 3th and 4th steps are repeated 20-40 cycles.

Denaturation step occure between 94 and 98°C.The heat breaks the hydrogen bonds and then causes the separation of the double strand of DNA into two single strands.
Annealing step occure between 50 and 65 °C. Primers anneals to DNA simple strand by complementarity of the nitrogenized bases.
Elongation step occure between 70 and 80°C, depending on the DNA polymerase used. The polymerase synthesizes a new DNA strand complementary to the DNA template

Preparation

Optimisation

Number of primer

Temperature

Gel electrophoresis analysis

Principle

Electrophoresis is a method using electrical field to separate DNA or RNA sequence by size. Smaller the fragment is, the more it migrates on the gel.
Using a DNA ladder, we can know the size of DNA sequence and then check if we have the sequence we wanted.

Preparation

Analysis