Reference gene selection for real-time quantitative PCR analysis of the mouse uterus in the peri-implantation period

PLoS One. 2013 Apr 24;8(4):e62462. doi: 10.1371/journal.pone.0062462. Print 2013.

Abstract

The study of uterine gene expression patterns is valuable for understanding the biological and molecular mechanisms that occur during embryo implantation. Real-time quantitative RT-PCR (qRT-PCR) is an extremely sensitive technique that allows for the precise quantification of mRNA abundance; however, selecting stable reference genes suitable for the normalization of qRT-PCR data is required to avoid the misinterpretation of experimental results and erroneous analyses. This study employs several mouse models, including an early pregnancy, a pseudopregnancy, a delayed implantation and activation, an artificial decidualization and a hormonal treatment model; ten candidate reference genes (PPIA, RPLP0, HPRT1, GAPDH, ACTB, TBP, B2M, 18S, UBC and TUBA) that are found in uterine tissues were assessed for their suitability as internal controls for relative qRT-PCR quantification. GeNorm(PLUS), NormFinder, and BestKeeper were used to evaluate these candidate reference genes, and all of these methods identified RPLP0 and GAPDH as the most stable candidates and B2M and 18S as the least stable candidates. However, when the different models were analyzed separately, the reference genes exhibited some variation in their expression levels.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Computational Biology / methods
  • Embryo Implantation / genetics*
  • Female
  • Gene Expression Profiling*
  • Gene Expression Regulation*
  • Male
  • Mice
  • Pregnancy
  • Uterus / metabolism*

Grants and funding

The research was granted by Scientific Research Foundation for Doctor (2010BSJJ012), Specialized Research Fund for the Doctoral Program of Higher Education (20110204120010), and Fundamental Research Funds for the Central Universities (QN2011105). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.