REFERENCE GENE SELECTION FOR QUANTITATIVE REAL-TIME PCR NORMALIZATION IN MOINA MACROCOPA EXPOSED TO PHENOL

To search suitable reference genes for normalization of quantitative Real-Time PCR (qRT-PCR) in Moina macrocopa, we tested three reference genes of β-actin, 16S rRNA and 12S rRNA by using four analysis methods: (1) expression level of the genes (cycle threshold value); (2) GeNorm; (3) NormFinder; and (4) BestKeeper. The results showed that the Ct values of the β-actin, 16S rRNA and 12S rRNA genes remained unchanged in M. macrocopa treated with different concentrations of phenol, and the order of the stability was 16S rRNA>12S rRNA>β-actin. GeNorm analysis revealed that the order of the stability was 16S rRNA=β-actin>12S rRNA. Both NormFinder and Bestkeeper software analysis demonstrated that the order of the stability was 16S rRNA>β-actin>12S rRNA. These results indicated that 16S rRNA was the best-fit reference gene for qRT-PCR in M. macrocopa, at least under phenol treatment, which provide useful information for future functional investigations of target gene expressions in M. macrocopa in response to environmental stress.