[关键词]
[摘要]
目的 对黄独微型块茎鲨烯合酶(SQS)基因的表达量进行分析, 旨在为阐明黄独微型块茎不同诱导形成时期皂苷的合成机制提供理论依据。方法 以Actin基因为内参基因, 采用实时荧光定量 PCR (qRT-PCR)分析技术。通过对不同诱导形成时期的黄独微型块茎进行取样并提取RNA, 反转录获得cDNA作模板, 利用sybr Green I成功构建了目的基因SQS和Actin的扩增曲线和熔解曲线。结果 定量结果表明, SQS基因在黄独微型块茎诱导形成的初期(第18~36天)。表达量显著增加;在黄独微型块茎诱导形成的中期(第36~72天)SQS基因的表达量显著下降, 并趋于稳定;在黄独微型块茎诱导形成的后期(第72~90天)SQS基因的表达量再次显著下降。结论 在黄独微型块茎诱导形成的过程中, SQS基因的表达量呈现出"低-高-低-不变-低"的变化趋势, 推测SQS是黄独微型块茎诱导形成中皂苷生物合成的关键酶。
[Key word]
[Abstract]
Objective In order to provide a theoretical basis for clarifying the mechanism of synthesis of saponins during different induction formation period of Dioscorea bulbifera microtubers, squalene synthase (SQS) gene expression of D. bulbifera microtubers was analyzed in this paper. Methods Using Actin gene as a reference gene, Real-time quantitative PCR (qRT-PCR) analysis technology was applied. The amplification curve and solubility curve of SQS (target gene) and Actin (reference gene) were successfully constructed by SYBR Green I after RNA was extracted from different formation period of D. bulbifera microtuber and cDNA was obtained by reverse transcription. Result Quantitative results showed that: During the initial stage (18—36 d) of D. bulbifera microtuber formation, the expression level of SQS gene increased significantly. During the mid-term (36—72 d) of D. bulbifera microtuber formation, the expression level of SQS gene decreased significantly, and tended to be stable. During the later stage (72—90 d) of D. bulbifera microtuber formation, the expression level of SQS gene decreased significantly again. Conclusion In general, in the process of D. bulbifera microtuber formation, the expression level of SQS gene shows the trend of "low-high-low-constant-low", which indicates that SQS is a key enzyme of saponin biosynthesis during the different induction formation period of D. bulbifera microtubers.
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[基金项目]
国家自然科学基金资助项目(31360072);江西省教育厅2014年度科学技术研究一般项目(GJJ14713)