Selection and validation of a set of reliable reference genes for quantitative sod gene expression analysis in C. elegans.

BMC Molecular Biology
David HoogewijsJacques R Vanfleteren

Abstract

In the nematode Caenorhabditis elegans the conserved Ins/IGF-1 signaling pathway regulates many biological processes including life span, stress response, dauer diapause and metabolism. Detection of differentially expressed genes may contribute to a better understanding of the mechanism by which the Ins/IGF-1 signaling pathway regulates these processes. Appropriate normalization is an essential prerequisite for obtaining accurate and reproducible quantification of gene expression levels. The aim of this study was to establish a reliable set of reference genes for gene expression analysis in C. elegans. Real-time quantitative PCR was used to evaluate the expression stability of 12 candidate reference genes (act-1, ama-1, cdc-42, csq-1, eif-3.C, mdh-1, gpd-2, pmp-3, tba-1, Y45F10D.4, rgs-6 and unc-16) in wild-type, three Ins/IGF-1 pathway mutants, dauers and L3 stage larvae. After geNorm analysis, cdc-42, pmp-3 and Y45F10D.4 showed the most stable expression pattern and were used to normalize 5 sod expression levels. Significant differences in mRNA levels were observed for sod-1 and sod-3 in daf-2 relative to wild-type animals, whereas in dauers sod-1, sod-3, sod-4 and sod-5 are differentially expressed relative to third stage la...Continue Reading

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Citations

Mar 20, 2010·International Journal of Legal Medicine·Antje KoppelkammMarielle Heinrich
May 3, 2013·Environmental Science & Technology·Tiffany Y LinJennifer A Jay
Aug 9, 2013·Nature Communications·Louis R LapierreMalene Hansen
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Methods Mentioned

BETA
PCR
GTPase
electrophoresis

Software Mentioned

qBase
geNorm
MFOLD
Primer3
geNorm VBA
Excel
Rotor
Gene

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