过量表达cyp71av1和cpr基因提高青蒿中青蒿素的含量

›› 2008, Vol. 10 ›› Issue (3): 64-70.

过量表达cyp71av1和cpr基因提高青蒿中青蒿素的含量

景福远1,张凌1,李美芽1,唐克轩1, 2

（1．上海交通大学农业与生物学院植物生物技术研究中心,复旦-交大-诺丁汉植物生物
技术研发中心,上海 200240； 2.复旦大学生命科学学院遗传工程国家重点实验室,
复旦-交大-诺丁汉植物生物技术研发中心,上海 200433)

收稿日期:2008-05-05 出版日期:2008-06-15 发布日期:2009-07-29
通讯作者: 唐克轩,教授,博士,主要从事植物分子生物学和植物生物反应器等研究。Tel: 021-34205916; E-mail: kxtang1@yahoo.com,kxtang1@163.com
作者简介:景福远,硕士研究生,主要从事植物次生代谢工程研究。
基金资助:
国家973计划项目(2007CB108805）和上海市科委项目（055407060）资助。

Over-expressing cyp71av1 and cpr Genes Enhances
Artemisinin Content in Artemisia annua L.

JING Fu-yuan1, ZHANG Ling1, LI Mei-ya1, TANG Ke-xuan1, 2

(1. Plant Biotechnology Research Center, School of Agriculture and Biology, Fudan-SJTU-Nottingham Plant Biotechnology
R&D Center, Shanghai Jiao Tong University, Shanghai 200240|2. State Key Laboratory of Genetic Engineering,
School of Life Sciences, Fudan-SJTU-Nottingham Plant Biotechnology R&D Center, Fudan University, Shanghai 200433,China)

Received:2008-05-05 Online:2008-06-15 Published:2009-07-29

摘要/Abstract

摘要：

青蒿素是从青蒿植株地上部分提取的一种含有过氧桥结构的倍半萜内酯,是目前最有效的治疗疟疾的药物。为了提高青蒿中青蒿素的含量,根据

GenBank中青蒿紫穗槐二烯氧化酶（CYP71AV1）和细胞色素P450还原酶（CPR）的基因序列设计特异引物,从青蒿cDNA中扩增出了cyp71av1和cpr

基因片段,分别连上CaMV 35S启动子和NOS终止子,然后将cyp71av1和cpr基因的表达盒依次插入到植物表达载体pCAMBIA2300中,获得含有

cyp71av1和cpr基因的植物表达载体p2300-GYR。通过农杆菌介导法转化青蒿,获得了20株转基因植株,PCR检测以及Southern杂交分析证实外源基

因已被成功地导入青蒿基因组中。采用HPLC-ELSD测定转基因青蒿中青蒿素的含量,结果表明,大部分转基因青蒿中青蒿素的含量高于对照植株,

其中,转基因青蒿中青蒿素含量最高约为对照的2.4倍。该研究证明过量表达cyp71av1和cpr基因是提高青蒿中青蒿素含量的有效方法。

关键词: 青蒿；青蒿素； cyp71av1基因； cpr基因

Abstract:

Artemisinin, a sesquiterpene lactone endoperoxide derived from plant Artemisia annua L., is the most effective anti-malarial

drug. In an effort to increase the artemisinin production, cyp71av1 and cpr genes were cloned from A. annua cDNA using two

pairs of gene-specific primers which were designed according to the gene sequences at GenBank. Both genes were inserted

between cauliflower mosaic virus (CaMV) 35S promoter and NOS terminator to construct expression cassettes, which were then

cloned into plant expression vector pCAMBIA2300, resulting in p2300-GYR containing both cyp71av1 and cpr genes.

Agrobacterium-mediated method was used to transform A. annua, and 20 independent transgenic plants were obtained. PCR

detection and Southern blot analysis demonstrated that the foreign genes were successfully introduced into A. annua genome.

Artemisinin contents in transgenic plants were determined by HPLC-ELSD and the result showed that artemisinin contents in

most transgenic plants were higher than that in control plants, with the highest artemisinin content in transgenic plant

being about 2.4-fold to that in the control plant. This study demonstrates that over-expressing cyp71av1 and cpr genes is an

effective approach to enhance artemisinin content in Artemisia annua L.

Key words: Artemisia annua L., artemisininin, cyp71av1 gene, cpr gene

中图分类号:

Q786

景福远1,张凌1,李美芽1,唐克轩1, 2. 过量表达cyp71av1和cpr基因提高青蒿中青蒿素的含量[J]. , 2008, 10(3): 64-70.

JING Fu-yuan1, ZHANG Ling1, LI Mei-ya1, TANG Ke-xuan1, 2. Over-expressing cyp71av1 and cpr Genes Enhances
Artemisinin Content in Artemisia annua L.[J]. , 2008, 10(3): 64-70.

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