Im having trouble with my qPCR assays. It seems like I'm getting a inhibitor in some of my samples when I'm producing cDNA. I use Invitrogen kits to DNase treat my samples then produce cDNA and sometimes after this the samples are fine, but other times the standard curves I make from these sample are completely off. The first few dilutions won't amplify properly and only by the 4th or 5th dilution do I get a normal sigmoidal curve. Its not due to inhibition in the original sample because I have produced cDNA from the same RNA sample and once its been fine, but the second time it wasn't. I can't work out what is causing this because I'm only adding reagents from the invitrogen kits. Does anyone have any idea of what could be going wrong?
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