I'm starting a new qPCR experiment and I would like to minimize the intra-plate and inter-plate variabilities, so I'm having several doubts.....
I've synthesised my cDNA (20 ul/diluted 1:5), but I've just realized that it won't be enough for all the genes that I want to analyse (30 genes in triplicate- 2ul each well). I know that it may sound silly, but I have two options but I'm not sure that I will avoid the RT variability:
1. do a second RT from the same patient and finish the last batch of genes with the new cDNA.
2. do RT from the same patient and mix the two together solutions together (old and new RT)
what do you suggest? Do you think that it will compromise my data?
Thanks a lot!