The Polymerase Chain Reaction (PCR) is a widely used biotechnology technique designed to amplify specific DNA sequences, and it operates through a cyclical series of three main steps.
The Three Key Steps of PCR
PCR efficiently creates millions of copies of a particular DNA segment by cycling through three temperature-dependent stages. These steps are performed repeatedly in a thermal cycler, a laboratory instrument that precisely controls temperature changes.
Here's a breakdown of the essential stages:
| Step | Description | Temperature (Approx.) | Purpose |
|---|---|---|---|
| 1. Denaturation | The double-stranded DNA template is heated to a high temperature, causing the hydrogen bonds between the base pairs to break. This separates the DNA into two individual single strands, making them available as templates. | 94-98°C | Separates DNA strands to expose templates for replication. |
| 2. Annealing | The reaction is cooled, allowing short, synthetic DNA sequences known as primers to bind (anneal) specifically to their complementary target sequences on each of the separated single DNA strands. These primers define the region of DNA to be amplified. | 50-65°C | Primers bind to define the region to be amplified. |
| 3. Extension | The temperature is raised again, and a heat-stable DNA polymerase (commonly Taq polymerase) extends the primers by adding free nucleotides (dNTPs) in the 5' to 3' direction. This synthesizes new, complementary DNA strands using the original strands as templates. | 70-75°C | Synthesizes new DNA strands. |
The PCR Cycle in Action
These three steps—denaturation, annealing, and extension—constitute one complete cycle of PCR. A typical PCR experiment involves 25 to 40 such cycles, leading to an exponential amplification of the target DNA. Each new DNA strand synthesized in one cycle can serve as a template in the subsequent cycles, resulting in the rapid generation of millions or even billions of copies of the desired DNA fragment from a tiny initial sample.
For more detailed information on the PCR process, you can refer to resources like Science Learning Hub.
