Screening of qPCR Internal Reference Genes in Stem Tissue of Pinus massoniana Under Hormone Treatment
QIN Hui-Juan1,2, FAN Fu-Hua1,2,*, ZHOU Zi-Jing1,2
1 Institute for Forest Resources and Environment of Guizhou/Key Laboratory of Forest Cultivation in Plateau Mountain of Guizhou Province, Guizhou University, Guiyang 550025, China; 2 College of Forestry, Guizhou University, Guiyang 550025, China
Abstract The utilization value of Pinus massoniana is extremely high, exogenous hormones can promote the secondary growth of its stem and wood formation, and rapidly increase its growth speed. Screening the best internal reference genes for qPCR experiment of P. massoniana stem under hormone treatment can provide technical support for gene expression research of secondary growth of P. massoniana. In this study, 14 candidate internal reference genes were selected from the stem transcriptome database of P. massoniana and gene expression was analyzed by qPCR. The expression stability under different hormone treatment - auxin (IAA), gibberellin (GA3), and brassinolide (BR) was analyzed by 3 softwares: GeNorm, NormFinder and BestKeeper. The selected internal reference genes were verified by the target gene cellulose synthase 2 (CESA2). The results showed that, under different hormone treatments, the internal reference genes with the best expression stability were phosphoate cytidine transferase 1 (PCYT1), ubiquitin-conjugating enzyme E2D(UBE2D), and phosphoglucomutase (PGM). The expression pattern of target gene CESA2 under hormone treatment confirmed the reliability of the above internal reference gene combination. The internal reference genes screened in this study would be helpful to improve the accuracy and reliability of qPCR experiment in P. massoniana, which provides a solid foundation for the analysis of differential gene expression of P. massoniana under hormone treatment.
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