携带pri-mir-20a的慢病毒表达载体的构建  

Construction of Pri-miR-20a Lentiviral Expression Vector

作　　者：马清华[1] 徐艳敏[1] 武佳[1] 李伟[1] 钱程[1] 刘立[1] 

机构地区：[1]浙江理工大学生命科学学院,杭州310018

出　　处：《浙江理工大学学报（自然科学版）》2010年第5期814-817,共4页Journal of Zhejiang Sci-Tech University(Natural Sciences)

基　　金：973计划(2004CB518804);浙江省"生物医学工程"重中之重学科开放基金(WYX0816)

摘　　要：采用PCR扩增获得的pri-mir-20a,过渡质粒pcDNA3.0-H1经BamHⅠ与HindⅢ双酶切后将两者连接产生pcDNA3.0-H1-pri-mir-20a重组质粒,再将PCR获得的H1-pri-mir-20a克隆到慢病毒载体pdc-SEW上获得重组慢病毒载体pdc H1-pri-mir-20a-SEW。将pCMV 8.91、pMD2.VSV.G、pdc H1-pri-mir-20a-SEW三质粒共转染293FT细胞,结果显示48 h后绿色荧光蛋白的表达达到80%,表明已成功构建出携带pri-mir-20a的慢病毒表达载体。该载体的成功构建为深入开展对mi R-20a的生物学功能和靶标验证提供了有效的工具。In this study,the pri-mir-20a amplified by PCR is inserted into the BamHⅠ/HindⅢ digested pcDNA3.0-H1 to generate recombinant pcDNA3.0-H1-pri-mir-20a.H1-pri-mir-20a is then cloned into lentiviral expression vector pdc-SEW to generate pdcH1-pri-mir-20a-SEW.Finally,pCMV 8.91,pMD2.VSV.G,pdcH1-pri-mir-20a-SEW are co-transfected cells of 293FT and the expression of green fluorescent protein reaches 80% in 48h.The result shows that the successful construction of lentivirus vector pdcH1-pri-mir-20a-SEW provides the basis for further study of biological functions and the targets of miR-20a.

关 键 词：pri-mir-20a 慢病毒载体 pdcH1-pri-mir-20a-SEW 

分 类 号：Q789[生物学—分子生物学]