肺炎球菌溶血素基因的克隆  

Cloning of the pneumolysin gene

作　　者：陈兵[1] 戴文佳[1] 王正敏[1] 李忠明 

机构地区：[1]复旦大学附属眼耳鼻喉科医院耳鼻喉科,上海200031 [2]海规生物科技(上海)有限公司,上海200436

出　　处：《中国耳鼻咽喉头颈外科》2006年第4期237-239,共3页Chinese Archives of Otolaryngology-Head and Neck Surgery

基　　金：国家自然科学基金(30471872);教育部211工程学科基金联合资助项目

摘　　要：目的应用基因工程技术克隆肺炎球菌溶血素(pneumolysin,Pn)基因,为研制中耳炎蛋白多糖结合疫苗奠定基础。方法根据Walker等肺炎球菌溶血素基因序列(GenBankaccessionno.X52474),设计一对各带有一个限制性酶切位点的引物,运用PCR从肺炎球菌的染色体DNA扩增出肺炎球菌溶血素基因。对该PCR产物进行酶切、回收,并将其克隆入表达载体pET-28a,然后转化至大肠杆菌JM109(DE3)宿主细胞中。结果通过常规基因克隆方法和PCR技术,成功构建重组体,酶切鉴定和基因测序证实构建正确。结论利用基因工程技术成功克隆了肺炎球菌溶血素基因。OBJECTIVE To prepare pneumolysin （Pn）by genetic engineering and thereby establish the basis for the study of vaccines against otitis media. METHODS A pair of primers including two restriction sites was designed based on the pneumolysin gene sequence reported by Walker in 1987. The pneumolysin gene was PCR-amplified from pneumococcal DNA. The resulting fragment, digested by restriction enzymes, was ligated into the vector PET-28a and then transformed into host cell E.coli JM109（DE3）. RESULTS The sequence of the inserted pneumolysin gene was confirmed by DNA sequencing. CONCLUSION The pneumolysin gene was successfully cloned into the host cell.

关 键 词：疫苗 中耳炎 链球菌 肺炎 溶血素类 

分 类 号：R764.21[医药卫生—耳鼻咽喉科]