PCR: The Polymerase Chain Reaction

Overview

The polymerase chain reaction, or PCR, is a technique used to amplify DNA through thermocycling – cyles of temperature changes at fixed time intervals. Using a thermostable DNA polymerase, PCR can create numerous copies of DNA from DNA building blocks called dNTPs. There are three steps in PCR: denaturation, annealing, and elongation. Denaturation is the first step in the cycle and causes the DNA to melt by disrupting hydrogen bonds between the bases resulting in single-stranded DNA. Annealing lowers the temperature enough to allow the binding of oligonucleotide primers to the DNA template. During the elongation step DNA polymerase will synthesize new double-stranded DNA.

This video provides an introduction to the PCR procedure. The basic principles of PCR are described as well as a step-by-step procedure for setting up a generalized PCR reaction. The video shows the necessary components for a PCR reaction, includes instruction for primer design, and provides helpful hints for ensuring successful PCR reactions.

Procedure

The polymerase chain reaction or PCR is a widely used method for amplifying DNA fragments. PCR uses thermocycling, which is the repeated heating and cooling of the reaction via three distinct temperatures called denaturation, annealing and extension or elongation.

The thermocycling reaction begins once the PCR reagents are put into a thermocycler a machine, which is programmed to precisely heat and cool the reaction.

The PCR cycle begins with denaturation, which occurs for 20 to 30