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

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

Abstract

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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Embryonic Stem Cells / metabolism
  • Gene Expression / genetics*
  • Mice
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • Reference Standards
  • Reproducibility of Results
  • Reverse Transcriptase Polymerase Chain Reaction / methods*
  • Reverse Transcriptase Polymerase Chain Reaction / standards

Substances

  • RNA, Messenger