Selection of reference genes for real-time RT-PCR normalization in brown alga Undaria pinnatifida

Quantitative real-time PCR (qRT-PCR) has become widely used method for detecting gene expression of an organism at different developmental stages or under various stressful environmental conditions. Finding reference genes is a prerequisite in this process. In this research, we investigated the expression stability of eight candidate genes in the brown alga Undaria pinnatifida, including EF1α (elongation factor 1-alpha), eEF1β (eukaryotic elongation factor-1 B beta), RPL35 (60S ribosomal protein L35-3), GAPDH (glyceraldehyde-3-phosphatedehydrogenase), RPS17 (40S ribosomal protein S17), LHCP (light-harvesting complex protein), ribH (6,7-dimethyl-8-ribityllumazine synthase), and ilvC (ketol-acid reductoisomerase). The haploid male and female gametophytes at different stages, as well as the diploid sporophytes treated with different abiotic factors were analyzed by geNorm and NormFinder software. It was found that eEF1β and ribH could be used as the most stable reference genes in studies of gender or developmental gene expression in the gametophytes. As for the detection of gene expression in the sporophytes, eEF1β and ribH were recommended as the most suitable reference genes for irradiance treatment, eEF1β and RPL35 for temperature treatment, while EF1α and LHCP for nutrition treatment. These results will enable us to go further for quantifying gene expression study at transcript level in U. pinnatifida.