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Team:LMU-Munich/Notebook/Protocols/25 PCR with Taq
From 2010.igem.org
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| - | Source: | + | Source: Fermentas Usage Information for the Taq DNA Polymerase (native) |
Latest revision as of 10:45, 29 September 2010
Source: Fermentas Usage Information for the Taq DNA Polymerase (native)
PCR with Taq Polymerase
In a sterile, nuclease free PCR-tube mix following components for 50µl:
Component
Volume/Final concentration
Taq DNA 10x Buffer (without MgCl2)
5 µl
dNTP 0.2mM each
5µl
upstream primer
0,1- 1 µM
downstream primer
0,1- 1 µM
DNA Template
10pg-1µg
Taq DNA Polymeras(3u/µl)
1,25u
nuclease free water to final volume of
50 µl
Recommended thermal cycling conditions for Taq Polymerase:
Step
Temperature
Time
Number of Cycles
Initial Denaturation
95°C
1-3min
1
Denaturation
95°C
0,5min
Annealing
42-65°C (dependent on primer and template)
30sec
25-40
Extension
72°C
1min (1min/kb)
Final Extension
72°C
5-15 min
1
Soak (end)
4°C (on our thermalcyclers 12°C)
Indefinite
1
