PCR – Polymerase Chain Reaction

A method of amplifying small amounts of DNA using the

principles of DNA replication

DNA Replication

Normal DNA replication requires the following: DNA gyrase DNA helicase SSBs DNA polymerase III andI DNA ligase RNA primers and primase Many nucleotides and template DNA

Polymerase Chain Reaction

PCR only requires: Single strand DNA

fragment Polymerase Primer Heat provides the

mechanism for H bonds to release, replacing gyrase, helicase, SSBs and primase

DNA primers replace RNA primers

PCR Machine

Taq Polymerase

Taq polymerases comes from a bacteria that lives in themrmal hotsprings and can tolerate the high heat of PCR (9r degrees C)

These bacteria were discovered in 1966, living in Yellowstone hot springs

The heat tolerant bacteria is essential to PCR

PCR Process

DNA sample is heated to 95 degreesDNA denatures, H bonds breakSample is cooled to 60 degreesDNA primers anneal to complementary

sectionsTaq polymerase enzymes build a

complementary strandDNA sample is heated to 95 degrees…

…2 hours later…

In 32 cycles at 100% efficiency, 1.07 billion copies of targeted DNA region are created.

20 cycles takes about 1 hourAmplified sample can be used in DNA

fingerprinting, sequencing, recombinant DNA, etc…

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