鞘蕊苏贝壳杉烯酸氧化酶基因的克隆及表达载体的构建  

Cloning of Coleus Ent-kaurenoic Acid Oxidase Gene and Construction of Expression Vector

作　　者：马朝芝 黄晓 余坤 尤朋涛 刘焱文 但汉雄 Ma Chaozhi;Huang Xiao;Yu Kun;You Pengtao;Liu Yanwen;Dan Hanxiong(College of Pharmacy,Hubei University of Chinese Medicine,Wuhan,43006)

机构地区：[1]湖北中医药大学药学院

出　　处：《基因组学与应用生物学》2018年第9期3919-3925,共7页Genomics and Applied Biology

基　　金：国家自然科学基金(31270405)资助

摘　　要：为提高鞘蕊苏有效成分的含量,开展Coleus Ent-kaurenoic acid oxidase（KAO）基因的克隆以及表达载体构建方面的研究。提取鞘蕊苏叶片的总RNA,利用套式PCR技术克隆获得KAO基因全长,将该基因连接到克隆载体p MD-18T,经测序鉴定正确后,再将该基因连接到p ET-28a表达载体中。结果显示,成功克隆出的鞘蕊苏KAO基因大小为1 500 bp,对克隆基因进行测序及酶切鉴定,均得到正确大小的DNA片段,说明表达载体构建成功。本研究成功克隆得到鞘蕊苏贝壳杉烯酸氧化酶基因并构建其表达载体,为提高鞘蕊苏有效成分含量提供了基础。In order to improve the content of effective components of Coleus, carried out the research on the cloning of the KAO gene of Coleus Ent-kaurenoic acid oxidase and the construction of expression vectors. The total RNA were extracted from the leaf blade of Coleus, and the full-length of KAO gene was obtained through the nested PCR technique, the gene was connected the cloning vector of p MD-18 T, then the gene was connected to the p ET-28α expression vector after sequencing identification. Result displayed that the size of the KAO gene was 1 500 bp, after the sequencing of cloning gene and enzyme-cutting identification, getting all the expected size of the fragment, proved that the expression vectors build success. This study successfully cloned the KAO gene and constructed the expression vector of the Coleus Ent-kaurenoic acid oxidase, which provided foundation for improving the effective ingredient content of Coleus.

关 键 词：鞘蕊苏 贝壳杉烯酸氧化酶 PCR 表达载体 

分 类 号：S567.239[农业科学—中草药栽培]