miR-610的克隆及其慢病毒表达载体的构建  

Cloning of has-miR-610 and Construction of Its Lentiviral Expression Vector

作　　者：张京伟[1] 王静[2,3] 李明梅[1] 石文韬[2] 丁琼[2] 魏蕾[2] 

机构地区：[1]武汉大学中南医院,湖北武汉430071 [2]武汉大学基础医学院,湖北武汉430071 [3]武汉市第一医院,湖北武汉430022

出　　处：《武汉大学学报（医学版）》2012年第5期663-666,共4页Medical Journal of Wuhan University

基　　金：国家自然科学基金资助项目(编号:30971132)

摘　　要：目的:克隆微小RNA hsa-miR-610并构建其慢病毒表达载体。方法:将PCR扩增得到的miR-610前体序列和pcDNA6.2-GW/EmGFP载体经双酶切后连接产生pcDNA6.2-miR-610真核表达载体,双酶切后测序鉴定,筛选阳性克隆。通过BP反应及LR反应将pre-miR-610克隆至慢病毒目的载体pLenti6/V5-DEST,构建miR-610的慢病毒表达载体pLenti6/V5-miR-610。结果:将构建好的慢病毒表达载体导入大肠杆菌Stbl3进行扩增,测序表明所构建的载体与预期的完全一致。结论:成功构建了miR-610的慢病毒表达载体pLenti6/V5-miR-610,为深入研究miR-610的生物学功能奠定了基础。Objective： To construct a lentiviral expression vector for hsa-miR-610.Methods： The pre-miR-610 amplified by PCR was inserted into pcDNA6.2-GW/EmGFP.The recombinant plasmid pcDNA6.2-miR-610 was confirmed by restriction endonuclease analysis.Using Gateway cloning technology,the lentiviral expression vector miR-610 was constructed by BP and LR recombination reactions.The recombinant plasmid pLenti6/V5-miR-610 was confirmed by DNA sequencing.Results： DNA sequencing demonstrated that the lentiviral expression vector pLenti6/V5-miR-610 was constructed successfully.Conclusion： The successful construction of lentivirus vector pLenti6/V5-miR-610 provides the basis for further study of molecular functions of miR-610.

关 键 词：hsa-miR-610 微小RNA 慢病毒 

分 类 号：R392.12[医药卫生—免疫学]