I'm going to be running some Taqman assays on cDNA generated from RNA collected from various samples, and will end up running a bunch of plates (right now the setup is 1/2 a plate per sample, including controls etc.). Since our thermocycler can only handle one 96-well plate at a time, it has been suggested that I make up the mixes all together, aliquot them into the plates, then store the plates at 4°C (well wrapped in plastic or Parafilm) until ready to run. I would imagine that the plates would be in the fridge for just one day, at max overnight.
My question is about the polymerase - can I add it ahead of time, and just take the plate out of the fridge and pop it in the thermocycler, or should I add it just before use? I've been kind of out of the DNA/RNA field for over 10 years, and I seem to remember from before that polymerases could have residual activity even at 4°C. Is this still the case, or have they improved to only be active at 60+°C or whatever your temperature is?
Alternatively, is there another ingredient I should leave out until just before amplification? cDNA? Probes? Something else? Also, does the storage time make sense? Is overnight okay, or should I run them all the same day? Is over the weekend okay? I don't want this question to extend into a too-broad "How do I do qPCR?" question, but any tips you have would be very helpful. As far as I'm aware, all the reagents are from Life Technologies/Applied Biosystems.
The reason I ask is because it seems to me that it would be much more accurate and reproducible to prepare the master mix with polymerase already included, instead of adding 1 μL of polymerase to each well just before running the assay.