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J Integr Plant Biol, 2003, 45 (5): 608-613, Research Article
Molecular Cloning, Escherichia coli Expression and Genomic Organization ofSqualene Synthase Gene from Artemisia annua
LIU Yan, YE He-Chun, WANG Hong, LI Guo-Feng
doi:
Abstract

A 1 539 bp squalene synthase (AaSQS) cDNA was cloned from a high-yield Artemisia annua L. strain 001 by reverse transcription-polymerase chain reaction(RT-PCR). The amino acid sequence of AaSQS is 70%, 77%, 44% and 39% identical to that of squalene synthases from Arabidopsis thaliana, tobacco, human and yeast, respectively. The AaSQS genomic DNA has a complex organization containing 14 exons and 13 introns. Full-length or C-terminal truncated cDNA was subcloned into prokaryotic expression vector pET30a and the constructed plasmid was introduced to Escherichia coli strain BL21 (DE3) for induced overexpression. No squalene synthase protein with expected molecular mass was observed in E.coli containing the putative full-length squalene synthase cDNA, however, overexpression in E.coli was achieved by truncating 30 amino acids of hydrophobic region at the carboxy terminus.

青蒿鲨烯合酶基因的克隆、结构分析与大肠杆菌表达
刘彦叶和春! 王红李国凤
(中国科学院植物研究所光合作用与环境分子生理重点实验室,北京IQQQJ‘)

 摘要: 用RT-PCR方法从青蒿(Artemisia annua L.)中克隆了一个1 539 bp全长鲨烯合酶cDNA.青蒿鲨烯合酶氨基酸序列与拟南芥、烟草、人类、酵母鲨烯合酶的一致性分别为70%、77%、44%和39%.青蒿鲨烯合酶基因组DNA结构很复杂,包括14个外显子和13个内含子.全长的或C末端截短的鲨烯合酶cDNA被克隆进原核表达载体pET30a并在大肠杆菌(Escherichia coli) BL21(DE3)中诱导表达.但在含有全长的鲨烯合酶cDNA的大肠杆菌中并没有观察到预期大小的鲨烯合酶表达,而C末端截短疏水区30个氨基酸的鲨烯合酶可在大肠杆菌中过量表达.    

关键词: 青蒿;鲨烯合酶;大肠杆菌过量表达;基因结构;克隆

通讯作者。E-mail: <hcye @ ns.ibcas.ac.cn>。

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