PCR is the abbreviation of "Polymerase Chain Reaction". It was invented by Kary Mullis in 1983. It is one of the methods of genetic engineering used to produce multiple copies of DNA, other being the cloning. It is used to amplify specific nucleic acids.
PCR versus Cloning
Use of PCR is becoming common day by day, but still it has not replaced cloning due to many factors. One major factor is:
â€¢ PCR uses a small piece of DNA to be amplified.
But PCR has one advantage over cloning i.e.
â€¢ It takes less time as compared to cloning.
Elements in PCR reaction
â€¢ DNA polymerase III (taq polymerase)
â€¢ dNTPs (deoxyribonucleotide triphosphate)
â€¢ Target DNA(template DNA strand)
â€¢ Buffer (to maintain pH)
Steps involved in PCR
There are three steps involved in PCR which are carried out at different temperatures.
The given DNA template is denatured with high heat (Ì´ 95Â°C). At this temperature the two strands of DNA are separated.
In step 2 each DNA primer starts binding to its complementary base sequence on the template DNA. It takes about 0.5 to 2 minutes and the temperature is maintained between 50 to 65 Â°C.
During this step the DNA polymerase III starts creating a new strand of DNA complementary to the template strand after recognizing the primer (this then replaces the primer) in the direction of 5Â´â†’3Â´. The temperature during this step is 72Â°C. As the temperature is high, the DNA polymerase is denatured. To avoid denaturation, "TAQ POLYMERASE" is used which was discovered from bacteria "Thermus aquaticus" (found in the hot springs). The taq polymerase can withstand high temperature without denaturation.
Major factor affecting PCR's working
The major factor affecting PCR's working is contamination. Even the small contamination could affect its working.
This contamination can also become a hurdle in crime labs, and this can lead us away from the fact. Now a day's modern labs consider this problem and try to overcome this problem.
Applications of PCR
â€¢ Detection of infectious (viral) diseases.
â€¢ Detection of gene mutations.
â€¢ Uses in forensic labs.
â€¢ Used to study base sequencing of DNA from mummified humans.
â€¢ Used in DNA finger printing.
Future of PCR
Due to its increasing uses it is predicted that it will be used in ever clinics. We will be able to take PCR on the road and analyzing DNA on the spot. Larger pieces of DNA will also be copied. It will be commonly used for sexually transmitted diseases and drug screening and genetic testing of animal and plant hybrids. Hence, it will be making human's life much more easier.
About Author / Additional Info: