雷公藤单加氧酶基因全长克隆及表达分析  

Cloning and mRNA Expression Analysis of Monooxygenase Gene from Tripterygium wilfordii

作　　者：周家伟[1] 刘雨佳[1] 胡添源 张逸风[1] 高伟[1] 黄璐琦[2] Zhou Jiawei;Liu Yujia;Hu Tianyuan;Zhang Yifeng;Gao Wei;Huang Luqi(Key Laboratory of Collateral Diseases ,School of Traditional Chinese Medicine, Capital Medical University, Beijing 100069, China;State Key Laboratory of Dao-di Herbs,National Resource Center for Chinese Materia Medica, Chinese Academy of Chinese Medical Sciences ,Beijing 100700, China)

机构地区：[1]首都医科大学中医药学院中医络病研究北京市重点实验室,北京100069 [2]中国中医科学院中药资源中心道地药材国家重点实验室培育基地,北京100700

出　　处：《世界中医药》2018年第2期259-263,共5页World Chinese Medicine

基　　金："十三五"时期北京市属高校高水平教师队伍建设支持计划项目(CIT&TCD20170324);市教委科技+社科计划重点项目(KZ201710025022);国家自然科学基金面上项目(81773830)

摘　　要：目的:单加氧酶(Monooxygenase,MO)是一类通过在底物上加一个氧原子的加氧酶,在动植物体内参与多种代谢途径。本实验致力于从雷公藤转录组数据中克隆得到单加氧酶基因c DNA全长,并对克隆得到的MO基因进行初步的功能分析。方法:通过对雷公藤转录组数据中筛选得到MO基因片段设计特异性引物,克隆MO基因c DNA全长,通过生物信息学方法对得到的MO基因进行分析,主要包括多重序列比对,蛋白结构预测和构建进化树分析等。结果:根据分析结果Tw MO基因全长为2 193 bp,共编码507个氨基酸,等电点为8.84,相对分子质量为55.20 k Da,多重序列比对显示它与其他植物的MO基因具有较高的同源性。雷公藤悬浮细胞经外源茉莉酸甲酯(Me JA)诱导后,实时荧光定量结果显示,TwMO的表达量明显增加,并在48 h达到最高,提示Tw MO基因与雷公藤的次生代谢产物的生成有关。结论:本研究首次从雷公藤悬浮细胞中克隆得到MO基因,为进一步研究该基因的功能奠定基础。Objective： Monooxygenase（ MO） is a kind of enzymes that can incorporate one atom of oxygen into the substrates in many metabolic pathways. This study aimed to clone the full-length c DNA of a MO from the transcriptome data of Tripterygium wilfordii and analyze its function. Methods： The full-length c DNA of monooxygenase（ Tw MO） was cloned from the transcriptome data of Tripterygium wilfordii by designing specific primers. The Tw MO gene was then analyzed by a series of bioinformatics analysis.Results： It showed that the full length of Tw MO c DNA was 2193 bp encoding 507 amino acids. The theoretical isoelectric point was8. 84 and the molecular weight was about 55. 20 k Da. Sequence alignment and phylogenetic analysis demonstrated that Tw MO had relative close relationship to MO from other species of plants. The relative expression level of Tw MO after Me JA inducing was upregulated and highest at 48 h,which suggested its regulatory role in special metabolites biosynthesis. Conclusion： This work laid the foundation for further studying the function of this monooxygenase in Tripterygium wilfordii.

关 键 词：雷公藤 克隆 基因 单加氧酶 表达分析 

分 类 号：R284.1[医药卫生—中药学]