Do I have to run 2 housekeeping genes (18S and beta-actin) together in each run in a Real-time PCR?

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Nadine A. Gund

Hi, the first question: Why do you want to use 18S and beta-actin? Cheers, Nadine

Jo Vandesompele

When doing relative quantification, there is no need that your references genes are measured in the same plate (run) as your genes of interest. It is perfectly okay to measure gene of interest in plate 1, and measure reference gene 1 in plate 2, and reference gene 2 in plate 3 (see Figure 2 - sample maximization strategy in attached paper).

Article Hellemans J, Mortier GR, De Paepe A, Speleman F, Vandesompel...