Team:LMU-Munich/Notebook/Protocols/25 PCR with Taq


PCR with Taq Polymerase

Source: Fermentas Usage Information for the Taq DNA Polymerase (native)

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