《植物生理学报》 2017, 53(9): 1680-1686

茅苍术内参基因筛选及其在活性成分生物合成研究中的应用

桑晓华, 谷巍^*, 巢建国, 刘青芝, 蒋玲, 韩赟, 周乐章

南京中医药大学药学院, 南京210023

通信作者：谷；E-mail: guwei9926@126.com

摘　要：

本研究选取茅苍术转录组数据库中18S rRNA、β-Actin、EF-1α、GAPDH、UBQ1和UBQ2作为候选内参基因, 运用实时荧光定量PCR技术分析6种候选内参基因在正常和干旱胁迫条件下不同组织中的表达, 并用Delta CT、geNorm、Norm-Finder、BestKeeper及RefFinder等方法评价候选内参基因的表达稳定性。结果表明, UBQ2和EF-1α在正常条件下不同组织中的表达均较稳定, 在干旱胁迫下EF-1α的表达最稳定。在此基础上, 以UBQ2作为内参基因, 研究正常条件下茅苍术活性成分生物合成关键酶HMGR、FPPS基因在其开花初期不同组织中的表达特性, 结果显示花中HMGR、FPPS基因相对表达量均最高, 根茎次之, 茎和叶中相对表达量均较低。

关键词：茅苍术; 实时荧光定量PCR; 内参基因; 3-羟基-3-甲基戊二酸单酰辅酶A还原酶; 法尼基焦磷酸合酶

收稿：2017-02-22   修定：2017-08-22

资助：国家自然科学基金(81573520)、江苏省中药资源产业化过程协同创新中心(ZDXM-3-24)、江苏省中药优势学科II期建设(ysxk-2014)、“六大人才高峰”高层次人才项目(2012-YY-009)和江苏省“333高层次人才培养工程”。

Selection of reference genes of Atractylodes lancea and its application in biosynthesis of active ingredients

SANG Xiao-Hua, GU Wei^*, CHAO Jian-Guo, LIU Qing-Zhi, JIANG Ling, HAN Yun, ZHOU Yue-Zhang

College of Pharmacy, Nanjing University of Chinese Medicine, Nanjing 210023, China

Corresponding author: GU Wei; E-mail: guwei9926@126.com

Abstract:

In this study, 18S rRNA, β-Actin, EF-1α, GAPDH, UBQ1 and UBQ2 were selected as candidate reference genes from the transcriptome databases of Atractylodes lancea, and the expression of six candidate reference genes in different tissues of A. lancea were analyzed under normal stage and drought stress, respectively, then the expression stability of the candidate reference genes was analyzed by using five methods of Delta CT, geNorm, NormFinder, BestKeeper and RefFinder. The results showed that the expression of UBQ2 and EF-1α were stable under different physiological condition, while EF-1α displayed the most stable under drought stress. On this basis, UBQ2 were chosen as the reference gene to evaluate the expression characteristics of HMGR and FPPS, which were the key enzyme of the biosynthesis progress of active ingredients, during the early stage of anthesis. According to the results, they all had a highest expression in flowers and a lowest expression in stems and leaves.

Key words: Atractylodes lancea; real-time quantitative PCR; reference genes; 3-hydroxy-3-methylglutaryl coenzyme A (HMGR); Farnesyl pyrophosphate synthase (FPPS)