Transcript Powerpoint Presentation: Polymerase Chain Reaction

POLYMERASE CHAIN
REACTION
PCR
Features
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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 few
hours
© 2010 Paul Billiet ODWS
Image Credit: Taq DNA polymerase
Procedure
Mix:
DNA sample,
specific complementary primers,
DNA polymerase
 Heat DNA to 95°C,
denatures DNA
Sister strands separate
 Cool to 56°C
Primers bind to DNA
Bracket region to be copied
Stop sister strand reannealing
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© 2010 Paul Billiet ODWS
Procedure
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Warm to 73°C
Optimum temperature for DNA polymerase
New sister strands synthesised from the primers
Heat again to 95°C
Strands 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
© 2010 Paul Billiet ODWS
Problems
Contamination of sample
 Any source of DNA will do
(bacteria, technician…..)
 Sterile conditions needed.
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© 2010 Paul Billiet ODWS