Ct's of goi higher and lower than ct's hkg - (Mar/28/2011 )
I have trouble with my RNA expression data. The Ct's of my housekeeping gene (Beta-actin) are sometimes higer and sometimes lower than the ct's of my gene of interest (some samples have much more expression than others). Does anyone know how to do statistical data analysis on such data?
It's natural that your HKG stays the same and GOI varies. What variation (what Cts) are we talking about?
It's always best to have HKG near your GOI values in general, but sometimes this isn't possible for all samples. If the difference in expression is not extreme, you calculate it as usual. If it is, you can't really do much about it, so you calculate it the same way but maybe add some note about the extremety of values.
Anyway, you usually want to prove a difference in GOI between your samples, and with extreme values you prove it, it just may not be that precise, like who cares if there is 100 000 fold or 500 000 fold increase when there is in general marked increase and that's what matters. (but as I said this would apply for extreme differences in values, probably not even your case)