I am new to RT-PCR data analysis and I hope someone can clarify my confusions as follows,
Following the literature, delta delta Ct method seems to be a common practice. For reference, the delta delta Ct steps are explained here - Computing fold change values for RT-PCR
Given that background, here is my question,
I have RT-PCR data with ~800 miRNAs. I want to do comparison between two groups (disease and control). Each group has 30 samples. I did global mean normalisation. That is, I normalised each Ct values from a sample to mean of all miRNAs in that sample. Boxplot of the data showed that distribution of Ct values across samples were different. I did between sample quantile normalisation. Now, after that I am planning to perform either limma based moderated t-scores or Mann-Whitney U test.
What is the difference between the approach I described above and delta delta Ct method? The approach I described seems simple and more intuitive to me, but I am worried whether I am missing some important point why delta delta Ct method is preferred.
I will appreciate for any clarification !