Evaluation of real-time PCR endogenous control genes for analysis of gene expression in bovine endometrium

Walker, Caroline G., Meier, Susanne, Mitchell, Murray D., Roche, John R. and Littlejohn, Mathew (2009) Evaluation of real-time PCR endogenous control genes for analysis of gene expression in bovine endometrium. BMC Molecular Biology, 10 Article number 1471: 100-112. doi:10.1186/1471-2199-10-100

Author Walker, Caroline G.
Meier, Susanne
Mitchell, Murray D.
Roche, John R.
Littlejohn, Mathew
Title Evaluation of real-time PCR endogenous control genes for analysis of gene expression in bovine endometrium
Journal name BMC Molecular Biology   Check publisher's open access policy
ISSN 1471-2199
Publication date 2009-11-01
Sub-type Article (original research)
DOI 10.1186/1471-2199-10-100
Open Access Status DOI
Volume 10
Issue Article number 1471
Start page 100
End page 112
Total pages 12
Place of publication London, England
Publisher BioMed Central
Language eng
Subject 1103 Clinical Sciences
0601 Biochemistry and Cell Biology
Formatted abstract
Quantitative real-time PCR gene expression results are generally normalised using endogenous control genes. These reference genes should be expressed at a constant level across all sample groups in a study, and should not be influenced by study treatments or conditions. There has been no systematic investigation of endogenous control genes for bovine endometrium to date. The suitability of both commonly used and novel endogenous control genes was evaluated in this study, with the latter being selected from stably expressed transcripts identified through microarray analysis of bovine endometrium. Fifteen candidate endogenous control genes were assessed across different tissue subtypes in pregnant and cycling Holstein-Friesian dairy cows from two divergent genetic backgrounds.

The expression profiles of five commonly used endogenous control genes (GAPDH, PPIA, RPS9, RPS15A, and UXT) and 10 experimentally derived candidate endogenous control genes (SUZ12, C2ORF29, ZNF131, ACTR1A, HDAC1, SLC30A6, CNOT7, DNAJC17, BBS2, and RANBP10) were analysed across 44 samples to determine the most stably expressed gene. Gene stability was assessed using the statistical algorithms GeNorm and Normfinder. All genes presented with low overall variability (0.87 to 1.48% CV of Cq). However, when used to normalise a differentially expressed gene (oxytocin receptor - OXTR) in the samples, the reported relative gene expression levels were significantly affected by the control gene chosen. Based on the results of this analysis, SUZ12 is proposed as the most appropriate control gene for use in bovine endometrium during early pregnancy or the oestrus cycle.

This study establishes the suitability of novel endogenous control genes for comparing expression levels in endometrial tissues of pregnant and cycling bovines, and demonstrates the utility of microarray analysis as a method for identifying endogenous control gene candidates. © 2009 Walker et al; licensee BioMed Central Ltd.
Keyword Induced Transcriptome Changes
Housekeeping Genes
Q-Index Code C1
Q-Index Status Provisional Code

Document type: Journal Article
Sub-type: Article (original research)
Collection: UQ Centre for Clinical Research Publications
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Created: Thu, 26 Aug 2010, 13:19:33 EST