The choice of reference genes for assessing gene expression in sugarcane under salinity and drought stresses

Sugarcane (Saccharum spp. hybrids) is a world-wide cash crop for sugar and biofuel in tropical and subtropical regions and suffers serious losses in cane yield and sugar content under salinity and drought stresses. Although real-time quantitative PCR has a numerous advantage in the expression quantification of stress-related genes for the elaboration of the corresponding molecular mechanism in sugarcane, the variation happened across the process of gene expression quantification should be normalized and monitored by introducing one or several reference genes. To validate suitable reference genes or gene sets for sugarcane gene expression normalization, 13 candidate reference genes have been tested across 12 NaCl- and PEG-treated sugarcane samples for four sugarcane genotypes using four commonly used systematic statistical algorithms termed geNorm, BestKeeper, NormFinder and the deltaCt method. The results demonstrated that glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and eukaryotic elongation factor 1-alpha (eEF-1a) were identified as suitable reference genes for gene expression normalization under salinity/drought-treatment in sugarcane. Moreover, the expression analyses of SuSK and 6PGDH further validated that a combination of clathrin adaptor complex (CAC) and cullin (CUL) as reference should be better for gene expression normalization. These results can facilitate the future research on gene expression in sugarcane under salinity and drought stresses.