Demystifying PCR: How Does it Work?

Demystifying PCR: How Does it Work?


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  • Source: Microbioz India

  • Date: 18 Apr,2024

Do you know Polymerase Chain Reaction (PCR)? PCR is a great tool for amplifying any targeted DNA section. It is widely used in such fields as molecular biology, genetics, medical diagnostics, forensics among others. The polymerase chain reaction (PCR) that may be defined as an artificial replication of DNA occurring in cells within a tube serves the purpose of creating millions of copies of a selected sequence on DNA.

Here are the steps involved in PCR:


This first step includes heating the mixture to temperatures above 95°C. By doing this, it breaks hydrogen bonds between complementary bases and separates the double-stranded DNA template into two single strands through denaturation. As a result, two single-stranded DNAs form.


After denaturation, the temperature around 50-65°C is cooled down by the reaction mixture which allows short DNA sequences known as primers to bind or anneal with specific sections of each single-stranded DNA molecule. Primers are important because they define a starting point for DNA synthesis by DNA polymerase while being usually about 18 – 25 nucleotides long designed with complementary sides for target region flanking sequences.

Extension (Elongation):

Once primers have annealed at this stage, the temperature rises up to approximately 72°C which is an optimal temperature required by PCR DNA polymerase enzyme such as Taq polymerase that can withstand heat if need be during thermocycling. Then, making use of deoxyribonucleotide triphosphates (dNTPs), these enzymes extend each primer towards its three prime end till they create new polynucleotide chain that fits exactly to what was originally there. As soon as it gives rise to another double-stranded helix, PCR results in doubling amount desired.

Repeat cycles:

In summary therefore Denaturation, Annealing and Extension form one cycle of PCR. The process is usually repeated for 20-40 times depending on the amount of DNA required to be amplified in a reaction mixture. Finally the target DNA sequence becomes increased exponentially with every cycle doubling that occurs. Eventually, even if there was a very minute amounts of starting matter, the PCR will have created millions to billions of copies for the target DNA sequence.

PCR has transformed molecular biology and its applications include genetic testing, gene cloning, sequencing as well as forensic analysis. Its adaptability together with its high rate and sensitivity makes it an essential tool in modern biological research and diagnostics.

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