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DNA Replication and PCR
8
DNA Replication and PCR
An overview
Players
Helicase
Unwinds the helix
Primase
makes an RNA primer to start the process
DNA Polymerase
Adds complimentary bases to make a new strand.
Applications in PCR
(Polymerase Chain Reaction)
What's in the mix?
DNA to be copied
Nucleotides
Buffer(contains MgCl2)
Primers
DNA polymerase
DNA Amplification
Number of copies produced:
PCR Reaction
95 C denaturation 30 sec
55 C annealing 30 sec
72 C elongation 30 sec
Repeat 30-40 times(cycles)
https://www.youtube.com/watch?v=QwT-Tj89VLo
Extra details
Semiconservative
The final double stranded DNA has a new strand and an old strand.
Exonuclease
Removes RNA primers
5' to 3'
Leading strand runs 5' to 3'
Lagging strand runs 3' to 5'
DNA polymerase builds 5' to 3'
Small complimentary pieces of DNA made by DNA Polymerase on the lagging strand
Fun Facts:
PCR was first invented in 1983
Taq polymerase (from Thermus equaticus) discovered in 1976.