Abstract Abstract Quantitative Real-time PCR is popularly used to study gene transcription levels in plants. Validation of candi-date reference genes is the key step to ensure accurate quantification. Nitrogen element assimilation and utilization play a critical role in sorghum (Sorghum bicolor) yield. The study was to select the most reliable reference genes for quantitative Real-time PCR (qRT-PCR) analysis of target sorghum genes under low nitrogen stress. We chose 7 commonly used housekeeping genes including Actin, ubiqutin conjugating enzyme (UBQ), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), UBQ10, eukaryotic translation initiation factor 2B (EIF2B), glyceraldehyde-3-phosphate dehydrogenase (sbGAPDH) and eukaryotic translation initiation factor 4a (EIF4a) to systematically assess their expression levels in leaves and roots of sorghum under low nitrogen stress by qRT-PCR. The results of primers specificity analysis showed that all the primers were in line with the requirements of the stability screening. The consequences of qRT- PCR indicated that the stability and expression level of candidate genes varied in the different tissues and under different treatments. The expression level of GAPDH was the highest in sorghum leaves, and that of UBQ10 was the highest in roots. We also analyzed and evaluated the stability of candidate reference genes by geNorm software. The results showed that GAPDH and EIF4a were the most suitable reference genes for normalizing target gene expression in sorghum leaves under low nitrogen stress, whereas sbGAPDH and EIF4a were recommended as the optimum combination in roots. In conclusion, this study has provided the basic data for expression analysis of target sorghum genes under low nitrogen stress and reference data for similar studies of other plants.
MA Jian-Hua,XUN Yi,YU Yu-Guo, et al. Screening of Reference Genes for qRT-PCR Analysis in Sorghum (Sorghum bicolor) Under Low Nitrogen Stress [J]. , 2017, 25(5): 805-812.
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