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Protocols PCR Mutation
From 2008.igem.org
Set up two separate primer extension reactions
One for each top and bottom primer
Cocktail:
40ul H2O
5ul 10X Pfu Buffer
1ul dNTP (10mM)
1ul Pfu Polymerase
1ul of each Primer (10uM)
1ul Template
_____________
50ul
ThermoCycle Program
94*C.......30 sec
_____________
95*C.......30 sec
55*C.......60 sec
68*C.......2 min/kb
Run for a total of 4 cycles
Combine 25ul from each extension reaction above. Add 1ul Pfu Polymerase.
Incubate as above for a total of 18 more cycles.
