POLYMERASE CHAIN REACTION

PCR

© 2016 Paul Billiet ODWS

Features Amplifies small samples

of DNA into millions of copies

Can copy the DNA of a single cell

Uses DNA polymerase from hot spring bacteria (Thermus aquaticus)

Automated to produce millions of copies in a few hours.

Taq DNA polymerase

© 2016 Paul Billiet ODWS

Procedure Mix:

DNA sample, specific complementary primers, DNA polymerase

Heat DNA to 95°C, denatures DNASister strands separate

Cool to 56°C Primers bind to DNABracket region to be copiedStop sister strand reannealing.

© 2016 Paul Billiet ODWS

Procedure Warm to 73°C

Optimum temperature for DNA polymeraseNew sister strands synthesised from the primers

Heat again to 95°CStrands separate again

Cool to 56°C more primers bind Warm to 73°C DNA polymerase synthesises new

sister strands Repeat cycle 10 times > 1000 copies Repeat cycle 20 times > 1 million copies.

© 2016 Paul Billiet ODWS

Problems Contamination of sample Any source of DNA will do

(bacteria, technician…..) Sterile conditions needed.

© 2016 Paul Billiet ODWS