Endy pcr

PCR in the Endy Lab
We generally use Invitrogen's Taq-based Platinum HiFi PCR supermix for most of our reactions. Following the manual is straightforward. The manual is available here (PDF). Read it before you use the mix for the first time and keep it for reference. If the link is broken, simply Google "Platinum PCR SuperMix."

Briefly, a typical reaction is set up as follows:

1. set up pre-labeled reaction tubes on ice

2. add the following components: - 45µL Platinum PCR SuperMix (rock gently after thawing, quick spin before use) - 200nM final concentration of each primer (insert calculation short cut here) - template DNA (note: the total volume of primers and template can be 0.5 to 20µL)

3. make sure reaction tubes are properly capped before placing in thermocycler

4. perform PCR with an initial heating step at 94C for 2 minutes followed by 25-35 cycles of 30sec at 94C, 30sec at 55C and 1Kb/min at 72C