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Publication type: Journal Article
Document type: Full Paper

Year: 2013

Author(s): Meyer, FR; Grausgruber, H; Binter, C; Mair, GE; Guelly, C; Vogl, C; Steinborn, R

Title: Cross-platform microarray meta-analysis for the mouse jejunum selects novel reference genes with highly uniform levels of expression.

Source: PLoS One. 2013; 8(5):e63125



Authors Vetmeduni Vienna:

Binter Claudia
Mair Georg
Meyer Florian
Steinborn Ralf
Vogl Claus

Vetmed Research Units
Institute of Topographical Anatomy
Unit of Molecular Genetics
Unit of Veterinary Public Health and Epidemiology
VetCore


Project(s): Molecular biomarkers of canine and feline primary osteosarcoma


Abstract:
Reference genes (RGs) with uniform expression are used for normalization of reverse transcription quantitative PCR (RT-qPCR) data. Their optimization for a specific biological context, e.g. a specific tissue, has been increasingly considered. In this article, we compare RGs identified by expression data meta-analysis restricted to the context tissue, the jejunum of Mus musculus domesticus, i) to traditional RGs, ii) to expressed interspersed repeated DNA elements, and iii) to RGs identified by meta-analysis of expression data from diverse tissues and conditions. To select the set of candidate RGs, we developed a novel protocol for the cross-platform meta-analysis of microarray data. The expression stability of twenty-four putative RGs was analysed by RT-qPCR in at least 14 jejunum samples of the mouse strains C57Bl/6N, CD1, and OF1. Across strains, the levels of expression of the novel RGs Plekha7, Zfx, and Ube2v1 as well as of Oaz1 varied less than two-fold irrespective of genotype, sex or their combination. The gene set consisting of Plekha7 and Oaz1 showed superior expression stability analysed with the tool RefFinder. The novel RGs are functionally diverse. This facilitates expression studies over a wide range of conditions. The highly uniform expression of the optimized RGs in the jejunum points towards their involvement in tightly regulated pathways in this tissue. We also applied our novel protocol of cross-microarray platform meta-analysis to the identification of RGs in the duodenum, the ileum and the entire small intestine. The selection of RGs with improved expression stability in a specific biological context can reduce the number of RGs for the normalization step of RT-qPCR expression analysis, thus reducing the number of samples and experimental costs.

Keywords Pubmed: Algorithms
Animals
Female
Gene Expression Profiling/methods*
Gene Expression Profiling/standards
Gene Expression Profiling/statistics & numerical data
Gene Regulatory Networks
Jejunum/metabolism*
Kruppel-Like Transcription Factors/genetics
Male
Meta-Analysis as Topic
Mice
Mice, Inbred C57BL
Models, Genetic
Molecular Sequence Data
Oligonucleotide Array Sequence Analysis/methods*
Oligonucleotide Array Sequence Analysis/standards
Oligonucleotide Array Sequence Analysis/statistics & numerical data
Proteins/genetics
Reference Standards
Reverse Transcriptase Polymerase Chain Reaction/methods*
Reverse Transcriptase Polymerase Chain Reaction/standards
Reverse Transcriptase Polymerase Chain Reaction/statistics & numerical data
Sequence Analysis, DNA
Ubiquitin-Conjugating Enzymes/genetics


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