Posted 06 March 2013 - 01:23 AM
I assume you're using the delta-delta Ct method and calculating relative quantity (RQ). For the delta-delta Ct method you are essentially calculating the ratio of two genes, so your measured gene might give the same Ct for two samples, but the reference gene (GAPDH, 18S, etc.) might have given a different Ct, leading to a different ratio....hence different RQs. The only problem is that you really don't know if your gene of interest is changing or your reference gene is changing. Since you are a beginner, it is usually best to assume that most of the big differences can be classified as "noise" associated with technical problems. Repeat the experiment until you can generate similar results 3 times. When you publish your data, someone should be able to follow your exact protocol and generate similar data. If you randomly chose one result over another because it supports your hypothesis, then you're essentially just making up data.