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１６ 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.
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
关键词
嗜热菌
热休克蛋白
基因克隆
基因表达
纯化
archaen
heat shock protein
gene cloning
gene expression
purification
