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How to Normalize of RT-qPCR with several reference genes ? - (Mar/04/2010 )

I've performed a RT-qPCR analysis with one target gene and two reference genes. Now I would like to calculate the mean normalized expression (MNE') of my target gene (Muller et al. 2002) using a normalization by multiple HKG (Vandesompele et al. 2002). My question is what should I do to adjust the calculation with the PCR efficiencies of my references? Should I take the geometric mean of the PCR efficiencies of my two reference genes?

Thanks in advance for your help!



if you want to normalize to more than one reference gene and adjust for different efficiences you need a different approach. you can find the formulas here:

pubmed link
Attached File


Thank you for your quick answer, it was really helpful !