I've just done a qRT-PCR on samples that have been treated with different concentrations of miRNA and I'm trying to assess for gene knockdown. I'm quite thrown with my results. From the raw Ct values of my gene of interest, it doesn't look like there's been much knockdown (Ct values between untreated control and treated samples are about the same or slightly elevated for the latter), but once I've normalised my data to the three most stable housekeepers the results are drastically different. Where the raw Ct values showed no difference, there is now a difference between control and miRNA treated samples. Which is very scary in my head at the moment.
So my question is have I done something wrong? Or am I freaking out over nothing?
Normalised Ct values reflect raw Ct values?
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