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Anal Biochem. 2008 Aug 1;379(1):127-9. doi: 10.1016/j.ab.2008.04.036. Epub 2008 Apr 26.

Standardization of real-time PCR gene expression data from independent biological replicates.

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Del E. Webb Neuroscience, Aging, and Stem Cell Research Center, Burnham Institute for Medical Research, La Jolla, CA 92037, USA.


Gene expression analysis by quantitative reverse transcription PCR (qRT-PCR) allows accurate quantifications of messenger RNA (mRNA) levels over different samples. Corrective methods for different steps in the qRT-PCR reaction have been reported; however, statistical analysis and presentation of substantially variable biological repeats present problems and are often not meaningful, for example, in a biological system such as mouse embryonic stem cell differentiation. Based on a series of sequential corrections, including log transformation, mean centering, and autoscaling, we describe a robust and powerful standardization method that can be used on highly variable data sets to draw statistically reliable conclusions.

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