Polymerase chain reaction
The polymerase chain reaction (PCR) is an in vitro technique used to amplify a selected DNA fragment. It allows a large quantity of a particular DNA sequence to be produced from a very small starting sample.
The reaction is carried out in a thermal cycler, which repeatedly changes the temperature so that the conditions required for each stage are produced automatically.
What PCR requires
| Component | Role |
|---|---|
| Template DNA | Contains the region that is to be copied. |
| Primers | Short single-stranded DNA sequences with bases complementary to sequences at the ends of the region being amplified. |
| Taq DNA polymerase | Catalyses the formation of new DNA strands and remains functional during the high temperatures used in PCR. |
| Free DNA nucleotides | Provide the nucleotides from which complementary DNA strands are constructed. |
| Buffer | Provides suitable chemical conditions, including an appropriate pH, for the reaction. |
One PCR cycle
- Denaturation: heat separates the two strands of the DNA molecule.
- Annealing: cooling allows primers to bind to complementary sequences on the exposed DNA strands.
- Extension: the temperature is raised to conditions suitable for Taq DNA polymerase, which forms complementary strands using free nucleotides.
These three stages are repeated many times. Each completed cycle can double the amount of the selected DNA fragment.
Exam Tip: When describing PCR, give the stages in the correct order and link each temperature change to what happens to the DNA, primers or Taq polymerase.