Polymerase chain reaction- Mechanism and Application

The Polymerase chain reaction or PCR is a simple and easy method for the production of multiple number of copies of piece of DNA in vitro. This method synthesised large quantities of DNA fragment without the gene cloning.

This technique was developed by Kary Mullis in California in 1985. He also share Noble prize for chemistry in 1993. The mechanism and detail of Polymerase chain reaction has been described by Erlich. The Polymerase chain reaction can be understood with the three steps:

Denaturation or Melting of target DNA: The target DNA is heated at about 94 degrees Celsius for 15 second. It separate two complementary strands of DNA. Each separate complementary strand now acts as template.

Annealing of Primers: Two primer that are made up of Oligonucleotide are annealed or fixed at 3' end of each  DNA template. This step is done at low temprature about 68 degrees Celsius for 60 seconds. The primers are attached with both template of DNA through hydrogen bond.

Primer extension: In this step, Deoxynucleoside triphosphate and a thermo stable DNA polymerase are added along with Mg ions. The optimum temperature for this step is about 72 degree Celsius. The DNA Polymerase catalyse the polymerization of primers according to separate DNA template.

The DNA Polymerase that is needed for extension, must be functional at high temprature therefore term thermostable can be used. For this purpose, Taq Polymerase and vent Polymerase are used which are isolated from thermophilic  bacteria like Thermus aquatics and Thermococcus litoralis.

After completion of one Cycle, the targeted sequences on both strand are copied and four strand are produced. This cycle may repeat 50 times. 

Remember 👌👌one million copies of target DNA sequence may produced  after the 20 cycle and one billion copies may produced after 30 cycles.

Application of Polymerase chain reaction : This technique has wide range of application in field of molecular biology, medicines and biotechnology.

It is used to amplified of DNA and RNA. It is used to diagnose of disease like AIDS, Tuberculosis,  Hepatitis and other infectious disease.

This technique is used to detect genetic Disorders like sickle cell anemia  phenylketonuria and muscular dystrophy.

It is most applicable in forensic science where it is used in search of criminals through DNA finger printing.

It is also applied in diagnosis of plant disease. A large number of plant pathogen are also detected by Polymerase chain reaction. 

It has also been found useful in field of archeology. It has been used to clear the doubt for the Wooly mammoth and Dinosaurs.