Polymerase Chain Reaction Process

The polymerase chain reaction or known as (PCR) is a scientific technique used in molecular biology to amplify a specific DNA sequence.It can be used very quickly and efficiently to produce millions or billions of copies of single DNA sequence.

Polymerase chain reaction or PCR uses repeated cycles of heating and cooling to make the copies of specific DNA. High temperature is necessary to break weak hydrogen bond that binds the two stands of DNA together and as a result multiple copies of a specific DNA sequence can be obtained.

In order to carry out PCR successful, following components are required.

Target DNA molecule,

A pair of DNA primers,

Heat resistant DNA polymerase,

All four types of deoxynucleotide triphosphate (dNTPs)

Target

Denaturation ( DNA strand separation)

Annealing ( Hybridization of primers)

Extension ( DNA synthesis, elongation,replication)

Step One: This is the first step of denaturation in which the double stranded DNA is heated to about 95 degree Celsius for about 15 seconds. Temperature is increased to break down the hydrogen bond that binds the double strands of DNA together to split into two single stands of DNA.

Step two: Once the two strands of DNA are separated , Temperature is reduced to 54 degree Celsius to cool down the mixture solution. DNA primers will begin to bind the flanking sequence. DNA primers binds to the 3 end of one strand and another primer binds to the 3 end of the complementary strand.

Step three: Deoxynucleotide triphosphate (dNTPs) and temperature resistant polymers are added in this elongation step. The temperature is slightly increased to 72 degree Celsius as it is the optimal temperature of heat resistant DNA polymerase

After one cycle of PCR ,two copies of DNA strand are produced and after two cycles , four copies of DNA strand are produced and so on. It multiplies after each cycle and make number of copies of DNA strands from single DNA.

The end result is an exponential increase in the total number of DNA fragments that include the sequences between the PCR primers, which are finally represented at a theoretical abundance of 2n.

Almost 20-30 cycles can be carried out in one hour.

PCR applications are helpful in diagnosis of majority of hereditary or non heritable diseases, this technique is also applied in molecular biology and microbiology to identify certain bacteria and viruses that can cause serious health problems. Applications of PCR in forensic science are also widely used now a days because only small amount of DNA samples are needed to diagnose several medical conditions.

Qualitative PCR can of course be used to detect not only human genes but also genes of bacteria and viruses. One of the most important medical applications of the classical PCR method is therefore the detection of pathogens. Many viruses contain RNA rather than