How far advance can you make PCR mix? - I'm a first time poster. (Oct/17/2008 )
Hi, I'm working on RT-PCR, so I need to make RT mix, do RT, then make PCR mix, do PCR.
To save time to make 2 mixes 1.5 hours apart, I'd like to see if I can make RT mix and PCR mix at the same time.
If I make PCR mix and store it at 4 degree until RT is done (takes 1.5 hours), will it effect the result?
Very appreciate your input.
should be no problem (especially if you leave out the primers). just make sure you keep track of which premix is which.
i would advice you to not add in the polymerase/reverse-transcriptase enzyme until when u are really going to do the experiment.
i have a thing with enzyme . if they are not being used they should be in their glycerol solution and not chilling around in 4C. loss of activity and unwanted reaction might occur in the mix at 4 C who knows.
You can buy ready-made PCR-master mixes (without primers and template DNA), e.g. from promega. Therefore it should be no problem.