Team:TU Darmstadt/Notebook/Methods/PCR with Pfu polymerase
From 2014.igem.org
PCR with Pfu polymerase
Short explanation:
PCR (polymerase chain reaction) is a method for exponentially amplifying a fragment of DNA in vitro.
Equipment:
- Micropipettes with sterile tips
- PCR machine
- PCR tubes
Chemicals & consumables: 50 µL batch
- 18 µL Pfu Mastermix 9:25
- 100 ng = 1 µL DNA template
- 1 µL Forward primer (10 µM)
- 1 µL Reverse primer (10 µM)
- up to 50 µL ddH2O
Procedure:
Standard protocol for Pfu Polymerasep>
# | Temperature | Time | ||
1 | 95°C | 00:05:00 | ||
2 | 95°C | 00:00:20 | ||
3 | 55°C | 00:00:30 | ||
4 | 72°C | 2 min/kbp | ||
5 | GO TO 2 | REPEAT 30x | ||
6 | 72°C | 3 min/kbp | ||
7 | 4°C | HOLD |