I have done the real-time qPCR and have the Ct for my target gene and two reference genes, but I don't know how to integrate the two reference genes in the calculation of the relative expression of my target gene. Usually, I use the delta-delta method to calculate the RQ, with only one reference gene. Has anyone already used multiple reference gene?
Christian De Santis
Hi there, you can use the delta-delta Ct (same formula) and use the geometric mean of the two (or three, or four, ...) HK genes as a normalization factor. It's the same approach used by genorm for example: http://genomebiology.com/2002/3/7/research/0034
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Laetitia Virlouvet
Thank you for your answer. I finally used this method with good results. Thanks
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