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作 者 ：王莹莹, 刘玉, 姬妍茹, 张正海, 周广麒, 刘宇峰

期 刊 ：生物技术通报 2014年 0卷 5期 页码：137-141

关键词：四尾栅藻；DGAT1基因；克隆；表达载体构建；

Keywords:DGAT1 Gene, Clone Expression vector, Construct,

摘 要 ：利用RNAiso Plus试剂盒提取紫苏总RNA，以其为模板采用RT-PCR方法扩增出紫苏DGAT1基因的cDNA编码区序列并构建克隆载体pMD-DGAT1，再将克隆载体pMD-DGAT1和pBI121空载体进行Xba I和BamH I双酶切，连接目的片段构建表达载体pBI121-DGAT1。结果显示，克隆得到目的片段长度为1 657 bp，与GenBank中紫苏DGAT1基因序列的最大同源性为97%，所构建表达载体pBI121-DGAT1双酶切得到的两条片段长度与连接前一致。结果表明，成功转化四尾栅藻并得到表达，转化后四尾栅藻的油脂含量较转化前提高近1倍。

Abstract:Total RNA of Perilla frutescens was extracted by RNAiso Plus kit. It was used as a template for RT-PCR to clone cDNA coding region of DGAT1 gene of Perilla frutescens, and cloning vector pMD-DGAT1 was constructed. Cloning vector pMD-DGAT1 and empty vector pBI121 were double-enzyme digested with Xba I and BamH I. Then expression vector pBI121-DGAT1 was constructed by linking the fragments. Results showed that 1 657 bp fragment was obtained by cloning and has 97% base similarity to DGAT1 gene of GenBank. Expression vector pBI121-DGAT1 was double-enzyme digested into two fragments. The length of them was as long as the ones before linking. It was transformed and expressed in the Scenedesmus quadricanda successfully. The transformed Scenedesmus quadricanda approximately double the oil content as before.

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