淋病奈瑟菌孔蛋白B原核重组质粒的构建、表达与蛋白鉴定  

Construction of Neisseria gonorrhoeae Porin B Plasmid Recombinant and Its Expression in E.coli and Identification of Fusion Protein

作　　者：叶嗣颖[1] 廖芳[1] 宋启发[1] 崔斌[1] 熊萍[1] 

机构地区：[1]华中科技大学同济医学院基础医学院病原生物学系,武汉430030

出　　处：《华中科技大学学报（医学版）》2005年第5期529-531,535,共4页Acta Medicinae Universitatis Scientiae et Technologiae Huazhong

摘　　要：目的克隆和分析淋病奈瑟菌孔蛋白(porinⅠB,PⅠB)的基因序列并表达相应的蛋白质,为淋病奈瑟菌感染的检测和基因工程疫苗的研究与开发建立基础。方法PCR扩增出孔蛋白PⅠB DNA片段后构建出重组质粒pET-PⅠB,并经质粒酶切筛选、DNA测序和异丙基硫代-β-D-半乳糖苷(IPTG)诱导大肠埃希菌DE3表达蛋白予以证实。结果所得PⅠB核苷酸序列与GenBank(AF090801)公布的序列相比较,同源性达99.28%,推导氨基酸序列同源性为98.14%。SDS-PAGE检测到40 kD大小的融合蛋白。结论淋病奈瑟菌孔蛋白PⅠB原核表达质粒构建正确。Objective To clone porin 15 （PⅠ15） sequences of Neisseria gonorrhoeae and analyze the protein expression as basis of research in detection, prophylactic and therapeutic vaccine against the pathogen infection. Methods PⅠB DNA fragment was amplified by PCR and cloned into prokaryotic expression plasmid pET-28a（ ＋ ） to construct a recombinant pET-PⅠB, which was verified by restriction nuclease, DNA sequencing and protein PⅠB expression in E. coli DE3 induced by IPTG. Results DNA sequence analysis revealed that the nucleic acid sequence of PⅠB gene was in 99.28% of homology and the putative sequence of amino acids was in 98.14%, compared with that （AF090801） published in GenBank. A 40 kD fusion protein was detected by SDS-PAGE. Conclusion A plasmid recombinant encoding porin 15 of Neisseria gonorrhoeae has been successfully constructed and expressed in E. coll.

关 键 词：淋病奈瑟菌 孔蛋白B 原核表达质粒 基因表达 

分 类 号：R378.1[医药卫生—病原生物学]