关键词: 窄竹叶柴胡, 内参基因, 柴胡皂苷, 实时荧光定量PCR, 表达分析

Abstract: Bupleurum marginatum Wall. ex DC. var. stenophyllum (Wolff) Shan et Y. Li is a perennial shrubby herb of the genus Bupleurum,with high medicinal value as the root is known as Tibetan Bupleurum. In this experiment,real-time quantitative PCR technology was used,combined with the three different data analysis softwares of the geNorm,NormFinder and BestKeeper to screen out the genes that can be stably expressed in different organs of B. marginatum var. stenophyllum from the five commonly used candidate genes:Actin,α-tubulin,β-tubulin,Cyclophilin and EF-1α as the internal reference genes,and then the selected internal reference genes were used to analyze the key enzyme genes of saikosaponin by the relative expression levels of HMGR,IPPI,FPS,SS,β-AS in different organs of B. marginatum var. stenophyllum. The results showed that β-tubulin gene could be used as an internal reference gene for its stable expression in different organs of B. marginatum var. stenophyllum;different saikosaponin synthase genes were expressed differently in different organs of B. marginatum var. stenophyllum. Except for IPPI gene which had the highest expression in seeds. The expression levels of HMGR,FPS,SS and β-AS in roots were higher than those in other organs,and the expression levels of HMGR,FPS and SS in lateral roots were higher than those in main roots. Therefore,the expression levels of key enzyme genes for saponin synthesis in different parts of B. marginatum var. stenophyllum were different. The optimal internal reference gene is β-tubulin gene,which can provide a basis for molecular biology research of B. marginatum var. stenophyllum and this research could promote its rational development and wide utilization.

Key words: Bupleurum marginatum var. stenophyllum, Internal reference gene, Saikosaponin, Real-time quantitative PCR, Expression analysis