Dear all,
has anyone experience in storing already pipetted qPCR plates before running them in the cycler? We are using the SensiFAST SYBR No-ROX Kit by Bioline according to the manufacturers protocol on a Roche Light Cycler 480. Due to organizational reasons of a practical lab course it would be great to know if and at which conditions the prepared qPCRs samples can be stored (of course in the dark, but 4°C, on ice or frozen).
Thanks a lot!
Gertrud
Ive stored a plate overnight at 4*C with no problems.Just make sure the plastic lid over your plate is sealed very well to avoid evaporation. I've known some people to freeze their plates @-20 for a week and use that, but I'd be really worried about losing samples to sublimation. If youre using Taq pol, the freezing wont really effect it + any shearing of your template should be inconsequential if your primers amplify anything less than 500bp (100-200 is the recommended product length).
Also if you're using an anti-body inactivated polymerase its much more likely to give you unwanted amplification before you run your plate than a chemically inhibited polymerase, so if you get some aberrations Id try switching to a chemically inactivated master mix.
From my experience, the potential sublimation in the freezer can be a real issue, even with good sealing, and I now avoid it at all costs. In my case, I only stored samples, without the master mix, to avoid spontaneous amplification. I have never tried at 4°C.
As far as I know, you can store them @ 4 over night in the dark. I did that and everything was fine.
Dear all,
thanks a lot for your helpful answers! Now I am sure that the students experiments will not be messed up by storing for a few hours at 4°C.
you can store, but you could have reproducibility troubles when repeating freshly experiments :))
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