hii friends… As you our daily opretion to getting something new information for you. So as like that now we discussing technology which is generally used biology or chemistry field. So this is nothing but the PCR Technique. PCR means Polymerase chain reaction. It is generally used for amplification of gene. It is a vitro technique.
This technique was invented by Kary Mullis in 1983. When he was working in Cetus Corporation (Biotech company), California. And for inventing this he was awarded by Nobel prize in chemistry in 1993.
What is PCR Technique ?
It is a gene amplification process i.e. it process multiple copies of known DNA sequence containing a gene are obtained. From this we can generate a billions of copies of a particular DNA sequence in very short time. For processing this technique PCR machine is used where several steps require to amplification of genes where carried out.
What are the basic requirements for PCR ?
For processing this technique it has some requirements for multiply copies of DNA. For this technique we must have a DNA segment which we have to multiply and its length is about 100 to 35,000 bp. It also require some primers called as forward and reverse primers which are synthetic oligonucleotides of 17 to 30 nucleotides. They are having complementary sequence from desired DNA fragments. Then four types of nitrogen bases that is dATP, dGTP, dTTP & dCTP are required for it. Also Taqpolymerase enzyme which thermaly stable and which can bear a temperature up to 94°C. This enzyme is isolated from bacterium Thermus aquaticus. Which is generally found in hot springs.
What are the steps of PCR Technique ?
There are three steps required for amplification of gene. This are Heat Denaturation, Annealing and Polymerization.
In this process DNA is heated at nearly 91°C. Because of this high heat breaking of hydrogen bonds of DNA is takes place which result, we get two separate single stand DNA. Which will then help in synthesis a new DNA fragments.
Genarally in annealing the single standed DNA which formed during heat denaturation process. This DNA is paired with primers. Which are designed as per requirements. This step is carried out at 55°C.
This step consist of extension of newly forming DNA. In that temperature is rised to 72°C. And with the help of taqpolymerase enzyme nitrogen bases are added behind the primers on single stand DNA.
When there is completion of these three process we can say it complete his one cycle. And in PCR process these cycles carried out 28 to 30 times. Also processing one cycle it takes 3 to 5 minutes. As PCR generates continues newer DNA. For further generation of DNA previously generated DNA is used as template DNA. This entire process is carried out in an automated machine named as PCR mchine.
Application of PCR
This technique used in medical or biological research labs. For recombinant DNA technology or cloning, gene amplification, functional analysis of genes, genome testing, DNA based phylogeny, DNA fingerprinting, diagnosis of hereditary diseases, detection and diagnosis of infectious disease ,for in genomic library, for industrial purpose etc.
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