Polymerase Chain Reaction

In a test tube

In an artificial setting and repeatedly in a laboratory process

• When scientists wish to produce large quantities of a particular sequence of DNA from very small quantities 
• This is used in genetic fingerprinting for forensic analysis or to amplify small quantities of rare DNA for cloning purposes

• A mixture of double stranded DNA 
• DNA nucleotides 
• A heat resistant version of DNA polymerase
• Short lengths of single strand DNA known as primers

• A sequence before the region of interest on the strand of the template 
• Just after the region of interest on the other strand

They provide the starting sequence for DNA replication and define the region to be amplified

• The double stranded DNA is split into 2 single stranded templates by heating to 95^oC for 30 seconds 
• This breaks the hydrogen bonds between the bases

• The mixture is cooled to 40^oC for 30 seconds 
• This allows the primers to hydrogen bond to the complementary bases on the templates

• The mixture is heated to 72^oC for 2 minutes 
• Nucleotides with a complementary base hydrogen bond to the template 
• This is the optimum temperature for the DNA polymerase enzyme which joins DNA nucleotides together with a phosphodiester bond to build up a new complementary strand alongside the template strand

The mixture is once again heated to 95^oC to separate the newly formed strands and the process is repeated for another 25 cycles (the number of strands doubles after each cycle)

• There is no risk of contamination as only fragments with the correct sticky ends are inserted into the plasmid. With PCR any contaminating DNA could also be copied 
• Bacteria will also produce the protein that the gene codes for 
• Much more accurate in copying DNA

It is extremely rapid due to lack of culturing cell needed in vivo