Selection of Reference Genes by Quantitative Real-Time PCR for Ruditapes philippinarum Exposed to Nitrite

In order to well investigate the eco-toxicity of nitrite on molluscs, a suitable housekeeper gene was needed for further analysis to qualify the toxicity-related genes. In the present study, the clam Ruditapes philippinarum was used as the target mollusc and cDNAs templates were got from gill tissue RNA of R. philippinarum after nitrite exposure. Six candidate genes were assayed in terms of their expression stabilities by geNorm and NormFinder softwares, followed by a comprehensive weighting comparison. GeNorm analysis indicated that the stability order for the six genes were Actin>CyPA>EF1alpha>Ubi>18S>Tubu. However, a little different order was suggested by the Normfinder software, Actin>Ubi>CyPA>EF1alpha>18S>Tubu. After a comprehensive weighting comparison, the final stability order of the six genes was Actin>CyPA>Ubi>EF1alpha>18S>Tubu. So, the Actin gene was the most suitable reference gene to qualify target genes for their expression levels in the clam R. philippinarum after nitrite exposure.