关键词: 肝肿瘤;细胞株;热休克蛋白72;克隆;基因表达

    摘 要:目的 克隆人热休克蛋白72(HSP72)基因，原核表达并纯化蛋白产物，以探讨其生物学功能. 方法 从热休克的人肝癌细胞株SMMC-7721中，用RT-PCR方法扩增得到HSP72基因并克隆到原核表达载体pQE-30中，在大肠杆菌JM109中用IPTG诱导下表达的蛋白经FPLC梯度洗脱和Sephacryl S200凝胶过滤纯化，经SDS-PAGE电泳后转印到NC膜上，用anti-HSP72单抗作探针，进行Western blot鉴定. 结果 克隆的基因序列分析结果与有关报道一致，所构建的pQE-30HSP72载体在大肠杆菌中表达出与预期大小相符的Mr 为72000的蛋白，经鉴定确系HSP72蛋白. 结论 克隆了人HSP72基因，并对该基因进行了原核表达与纯化，为进一步研究其生物学功能奠定了良好基础.

    Cloning，prokaryotic expression and purification of human heat shock protein72gene

    YU Hou-You SONG Zu-Jun YUAN Shun-Shu LIU Yan-Fang YANG Shou-Jing

    1 Department of Emergency Medicine，Xijing Hospi-tal，

    2 Department of Pathology，Faculty of Preclini-cal Medicine，Fourth Military Medical University，Xi'an710033，China

    Keywords:liver neoplasms;cell line;heat shock protein72;cloning;gene expression

    Abstract:AIM To clone human heat shock protein72(HSP72)gene，do prokaryotic expression and purify HSP72protein for further study.METHODS Human HSP72gene was obtained by RT-PCR from heat shocked human hepato-carcinoma cell line SMMC-7721.Its product was recombined in vitro with prokaryotic expression vector pQE-30and was transformed to E.coli strain JM109.The proteins ......