Denise Yu
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Hot Start PCR (11/10/08)
Reagent
-10x Thermo-Start Standard Buffer (5 microliters)
-20mM dNTP Mix (2 microliters)
-25mM MgCl2 (3 microliters)
-Primer Mix (2.5 microliters)
-Template DNA 10 ng
-Thermo-Start DNA Polymerase (.25 microliters)
-double-distlled, sterile H2O (up to 50 microliters)
Thermal Cycle
Enzyme Activation
1. 95C for 15 minutes
Denaturation
2. 92-95C for 10-30 seconds
Annealing
3.50-65C for 30 seconds, repeart 25-30
Extension:
4.72C for 30 seconds
Final Extension:
5. 72C for 5 minutes