人肿瘤MUC1/Y基因真核表达载体的构建及在BIU-87细胞中的表达  

Construction of eukaryotic expression vector EGFP-MUC1/Y and its expression in bladder carcinoma cell line BIU-87

作　　者：罗刚[1] 周四维[1] 鲁雄兵[1] 胡志全[1] 宋晓东[1] 孙凯[1] 曹正国[1] 叶章群[1] 

机构地区：[1]华中科技大学同济医院泌尿外科,武汉430030

出　　处：《临床泌尿外科杂志》2005年第9期574-576,580,共4页Journal of Clinical Urology

基　　金：国家自然科学基金资助项目(No:30271300)

摘　　要：目的:克隆人肿瘤MUC1/Y全长基因及构建真核表达载体EGFP-MUC1/Y,并观察EGFP-MUC1/Y在BIU-87细胞中的表达,以进一步用于生物学功能及肿瘤生物学治疗的研究。方法:取新鲜膀胱肿瘤组织,提取总RNA,采用随机引物进行逆转录,将特异引物扩增的MUC1/Y全长PCR产物克隆至真核表达载体pEG-FP-C1,测序鉴定后命名为EGFP-MUC1/Y,用脂质体转染膀胱肿瘤细胞BIU-87,以Western Blot检测其表达情况。结果:酶切鉴定和序列分析证实构建质粒含人MUC1/Y全长cDNA编码序列,Western Blot检测和转染实验表明,构建的pEGFP-MUC1/Y基因在BIU-87真核细胞中表达,其绿色荧光蛋白瞬时表达率为20%。结论:人肿瘤MUC1/Y全长cDNA基因克隆及其真核表达载体pEGFP-MUC1/Y构建成功,可用于肿瘤生物学治疗的研究。Objective：To clone human full length MUC1/Y cDNA and construct an eukaryotic expression vec tor EGFP-MUC1/Y, then transfect it into bladder carcinoma cell line BIU-87 and detect its expression with Western Blot and green fluorescent protein, Methods：The total RNA of human bladder carcinoma was extracted and amplified by RT PCR primed by random and specific primers respectively. The amplified products were cloned and linked in green fluorescent protein vector EGFP-C1. The recombinant EGFP vector was transfect into human bladder tumor cell BIU-87 by Lipofectamine. Results：The cDNA sequencing showed that the recombinant plasmid contained the coding region of human full length MUC1/Y gene and could be expressed 20% of BIU-87 cells. Conclusions：The human full-length MUC1/Y gene was cloned and expressed in human bladder carcinoma cell line BIU- 87, our study indicated its potential roles in tumor gene therapy.

关 键 词：膀胱肿瘤 MUC1/Y 克隆 真核表达 

分 类 号：R737.14[医药卫生—肿瘤]