Screening the Reference Genes for Quantitative Gene Expression by RT-qPCR During SE Initial Dedifferentiation in Four Cultivars that Have Different SE Capability.

RNA sequencing (RNA-Seq)-based gene expression analysis is applicable to a wide range of biological purposes in various species. Reverse transcription quantitative PCR (RT-qPCR) is also used to assess target gene expression utilizing stably expressed reference genes as internal control under a given set of conditions. However, investigations of the reference genes for RT-qPCR normalization in the process of somatic embryogenesis (SE) initial dedifferentiation in are rarely reported. In this study, on the basis of our previous transcriptome data of three different induction stages during SE initial dedifferentiation process in four cultivars that have different SE capability, 15 candidate genes were selected during SE initial dedifferentiation process, and their expression stability was evaluated by geNorm, NormFinder, and BestKeeper. The results indicated that the two genes of () and () showed stable expression in the four different cultivars, endowing them to be appropriate reference genes during three induction stages in the four cotton cultivars. In addition, the stability and reliability of the two reference genes of and were further verified by comparing the expressions of () and () between RT-qPCR results and the RNA-seq data, which showed strong positive correlation coefficient (R = 0.8396-0.9984), validating again the steady expression of and as the reliable reference genes. Our results provide effective reference genes for RT-qPCR normalization during SE process in different cultivars.