Screening of reference genes in real-time PCR for .

Six candidate reference genes were chosen from the transcriptome database of using the bioinformatics method, including four conventional reference genes , Eukaryotic translation initiation factor 5A (), Tubulin alpha (, ubiquitin ()) and two new candidate reference genes (Ribosomal protein S21 () and Serine/threonine protein phosphatase PP1-β catalytic subunit (-PP1)). In addition, a traditional reference gene 18S ribosomal RNA () obtained from NCBI databases was also added to the analysis. Real-time PCR was used to detect the expression of seven candidate reference genes in six populations of and four developmental stages (female, male, larva and egg) of a population. The stability of the expression of candidate genes was evaluated by three software programs, BestKeeper, geNorm and NormFinder. The results showed that is the most suitable reference gene for gene functional research of different populations, while both and are the most suitable reference genes for different developmental stages of a population. Therefore, is the best reference gene for studying . However, one defect of this study is that only seven candidate reference genes were analyzed; ideally, more genes should be tested.