Identification of suitable reference genes of Scylla paramamosain for gene expression profiling in various tissues and under vibrio challenge

Abstract The quantitative real-time transcription-polymerase chain reaction (qRT-PCR) is now used widely in studies about mRNA expression levels. The selection of one or more stable reference gene(s) used for data normalization is substantial. In this study, expression levels of eleven candidate reference genes ( β-actin , 16S rRNA , 18S rRNA , 28S rRNA , α-I tubulin , GAPDH , ribosomal protein L13 , elongation factor 1 α , elongation factor 2 , arginine kinase and ubiquitin ) were examined using the GenomeLab GeXP analysis system (Beckman Coulter). Gene expression data were analysed using two different statistical models: geNorm and NormFinder . (1) In six different tissues (hepatopancreas, haemocytes, heart, gill, muscle, and testis) from the mud crab, Scylla paramamosain , 18S rRNA and elongation factor 1 α were identified as the two best reference genes. (2) In the haemocytes after being challenged by Vibro parahaemolyticus , the result suggested that ubiquitin was the most stable gene after the treatment. 18S rRNA , elongation factor 1 α and ubiquitin are herein recommended as the best combination. These results provide useful options for reference gene selection under different experimental conditions in qRT-PCR studies in the mud crab.