I've done my analysis on my Ct values. Firstly when I checked if there was downregulation or upregulation (I wanted downregulation as I was performing gene silencing) I used the Livak method, no problems. So for one experiment I got 0.166 so downregulation as I wanted, that's fine.
However, to be able to do statistical analysis and compare my results, I calculated my ratios (2^Ct values) for each sample separatley normalised to the controls of the target & reference gene (means) and got a value of 0.1828.
What I don't understand is why are these two values different? Am I not looking for the same thing which is the rate at which downregulation occured? And if so why is it two different values?
What I do follow is that when you're trying to find whether it's been silenced or not, you look at the overall values together and that confirms whether it has or not...my main question is what is the value in which it has been deregulated as I've one from Livak method using the means of Ct values and then another method which provides average ratios
Edited by J092, 21 February 2015 - 03:44 PM.