Polymerase chain reaction (PCR) is a powerful technique used in several areas of research. PCR takes advantage of the basic principles of DNA replication and allows for specific copying of a DNA sequence.
Here is a 3-D simulation of the process:
A PCR reaction includes 3 basic steps which are repeated for 20-30 cycles:
- Denature: The two strands of a DNA molecule are split from each other by simple heating. Recall that the two sides of a DNA helix are held together by relatively weak hydrogen bonds between the base pairs.
- Anneal: Each PCR reaction uses a specific pair of DNA primers (short, single stranded DNA molecules). These primers are designed to flank the area of DNA to be copied in the reaction. The reaction is cooled to a temperature (somewhere between 50-65° C) where the DNA strands start to reattach, and the primers stick to their complementary sequence.
- Extend: The PCR reaction is heated to 72°C which is the optimum temperature for a DNA polymerase enzyme called Taq polymerase. Taq polymerase is derived from the hot spring bacterium Thermus aquaticus and is able to remain stable through the denaturing step of each cycles. The Taq polymerase attaches to each primer and fills in the missing nucleotides in the 5’ to 3’ direction.
During each cycle of the three steps the amount of DNA present in the reaction
doubles.
Once you feel comfortable with the basic idea of PCR, read this technical chapter on PCR from Molecular Cloning: