Team:EPF-Lausanne/Protocols/1.5 steps PCR

From 2009.igem.org

1.5 Steps PCR Protocol




1. Do the following mixing:

dNTP 1 μl
5x Buffer + MgCl2 10 μl
Primer Mix 1 μl
template 0.5 μl
HiFi Plus DNAp 0.5 μl
dH2O 36.5 μl
Final Volume 50 μl


2. Make it run 10 cycles:

1             94°C 4:00
2             94°C 0:30
3             55°C 1:00
4             72°C 2:00
5             Cycle 2-4 10 times
6             72°C 7:00
7             4°C


3. We now add the standard iGEM primers which finish the extension, and do the following cycles:

1             94°C 4:00
2             94°C 0:30
3             55°C 1:00
4             72°C 2:00
5             Cycle 2-4 25-30 times
6             72°C 7:00
7             4°C