嗜热菌热休克蛋白基因的克隆与表达  

Cloning and expression of heat shock protein gene of a Hyperthermophilic bacteria in E.coli .

作　　者：颜真[1] 王俊楼[1] 韩苇[1] 赵永同[1] 张英起[1] 苏成芝[1] 

机构地区：[1]第四军医大学,陕西西安710033

出　　处：《第四军医大学学报》1999年第7期587-589,共3页Journal of the Fourth Military Medical University

基　　金：全军医药卫生科研基金

摘　　要：目的：克隆和在原核表达系统中表达超嗜热古球菌 Ｋ Ｏ Ｄ１（ｈｙｐｅｒｔｈｅｒｍ ｏｐｈｉｌｉｃ Ｐｙｒｏｃｏｃｃｕｓｓｐ． Ｋ Ｏ Ｄ１， Ｈ Ｐ Ｋ）之热休克蛋白基因β亚基（ｃｐｋ Ｂ）． 方法：用 Ｐ Ｃ Ｒ技术从 Ｈ Ｐ Ｋ 基因组 Ｄ Ｎ Ａ 中扩增ｃｐｋ Ｂ基因片段，经克隆和核苷酸序列分析后再克隆至原核表达载体ｐ Ｂ Ｖ２２０，升温诱导ｃｐｋ Ｂ基因表达，利用 Ｃｐｋ Ｂ蛋白的耐高温特性纯化该蛋白． 结果：扩增和克隆了１６ ｋｂ 的ｃｐｋ Ｂ基因，并经 Ｄ Ｎ Ａ 测序证实，实现了ｃｐｋ Ｂ基因在原核系统 Ｐ Ｌ Ｐ Ｒ 启动子控制下的表达，建立了纯化 Ｃｐｋ Ｂ蛋白的方法． 结论：该研究为进一步探讨 Ｃｐｋ Ｂ蛋白的结构与功能奠定了基础．AIM: To clone and to express β subunit of heat shock protein gene ( cpk B) of a hyperthermophilic pyrococcus sp . KOD1 (HPK) in E.coli . METHODS: Polymerase chain reaction (PCR) was employed to amplify the 1.6 kb cpk B gene fragment from HPK genome. The fragment was cloned, sequenced and then constructed into E.coli expression vector pBV220 with P LP R promotor. cpk B gene was expressed in E.coli by the induction of temperature shift from 32℃ to 42℃ in LB culture medium. The expressed CpkB protein was purified by heating treatment and ion exchange. RESULTS: The 1.6 kb cpk B gene was amplified, cloned, sequenced and expressed in E.coli . The purification method of the expressed CpkB protein was also constructed. CONCLUSION: The results of our research has laid a basis for further study of the stucture and function of CpkB protein.

关 键 词：嗜热菌 热休克蛋白 基因克隆 基因表达 纯化 

分 类 号：Q78[生物学—分子生物学]