Polymerase chain reaction is the process of gene amplification and gene isolation in artificial conditions

1. Polymerase Chain Reaction
(PCR)
By
Virendra Singh

2. What is PCR
• PCR is the process of DNA (gene of interest) amplification under in
vitro conditions.
• The ability of DNA polymerases is exploited to synthesize new
DNA strands
• Chemically synthesized oligonucleotide primers provide 3’ OH
required for extension of DNA in the presence of DNA polymerases.
• The copies of DNA produced by PCR reaction are also known as
Amplicons.
• This technique was invented by Kary Mullis in 1985. He got
Nobel Prize in chemistry for this invention.

3. Requirements of PCR reaction
 Target DNA or the gene to be amplified.
 Two primers (Forward Primer and Reverse primer)
 Taq DNA polymerase (DNA polymerases)
 Deoxynucleotide triphosphate ( dNTPs)
 MgCl2 , it acts as the cofactors for the DNA polymerases. It provides
divalent cation.

4. 1. Denaturation of DNA by heat at 94˚C
3’
5’ 3’
5’
5’ 3’
3’ 5’
3’ 5’
5’ 3’
3’
3’5’
5’
2. Primer annealing at 54˚C
3. Primer Extenstion at 72˚C
5’ 3’
3’ 5’
3’ 5’
3’5’
Steps 1, 2, 3 Repeat 25- 30 cycles
PCR Reaction Cycle

5. Applications of PCR
1. The traces of DNA can be amplified large amount for forensic
purposes
2. Purification of desired known gene from the length of DNA
Limitations of PCR
1. Only 5 kb of DNA can be easily amplified.
2. Gene of already known sequence can be isolated from DNA.