小鼠PMX-IRES-Wnt5a逆转录病毒载体的构建及在3T3细胞中的表达  

Construction of mouse PMX-IRES-Wnt5a retroviral expression vector and expression of Wnt-5a in NIH3T3 cells

作　　者：严春 刘彩霞[2] 闫绍荣 李强[1] 林森[1] 吴文兵[1] 吴礼峰[1] 陈炯[3] 

机构地区：[1]浙江省瑞安市人民医院检验科,浙江瑞安325200 [2]温州医学院附属第二医院检验科,浙江温州325027 [3]浙江省瑞安市人民医院烧伤科,浙江瑞安325200

出　　处：《中国现代医生》2012年第25期7-8,10,共3页China Modern Doctor

基　　金：浙江省温州市2009年第一期科技计划项目(Y20090097)

摘　　要：目的构建小鼠Wnt-5a逆转录病毒载体,观察Wnt-5a在3T3细胞的表达情况。方法采用同源重组技术将小鼠Wnt-5a插入逆转录病毒载体PMX-IRES-GFP,构建PMX-IRES-Wnt5a重组质粒,利用脂质体转染3T3细胞,RT-PCR检测Wnt-5a mRNA表达情况,Western-Bloting检测Wnt-5a蛋白表达情况。结果经限制性内切酶证实成功构建了携带Wnt-5a基因的逆转录病毒载体PMX-IRES-Wnt5a,RT-PCR检测结果显示Wnt-5a在转染的3T3细胞的表达明显增高。结论成功构建Wnt-5a逆转录病毒载体,并能在3T3细胞成功表达Wnt-5a。Objective To construct the retroviral expression vector of PMX-IRES-Wnt5a,transfect into 3T3 cells,and observe the expression of Wnt-5a（nethods Gene recombinant technology was employed to clone mouse Wnt-5a gene to the retroviral expression vector of PMpX-IRES-GFP, to obtain the recombined plasmid PMX-IRES-WntSa,transfect into 3T3 cells by Lipofectamine 2000. The result of transfection was analyzed by RT-PCR and Western-Blotting. Results Identifica tion by enzyme digestion confirmed successful construction of the retroviral expression vector of PMX-IRES-Wnt5a. RT-PCR revealed that the expression of Wnt-Sa mRNA and protein was obviously increased in 3T3 cells after transfection. Conclusion The retiroviral expression vector of PMX-IRES-WntSa is successfully constructed and stably expressed in 3T3 cells.

关 键 词：WNT-5A 逆转录病毒载体 

分 类 号：R782[医药卫生—口腔医学]