亚洲牛带绦虫Spef1-Like基因克隆、表达及纯化  

Cloning and expression of Spef1 like gene from Taenia asginata asiatica

作　　者：王杰[1] 戴佳琳[2] 黄江[2] 吴璇[2] 廖兴江[2] 申萍香[2] 周灵贵[2] 杜武英[2] 郎书源[2] 

机构地区：[1]贵阳医学院法医教研室,贵阳550004 [2]贵阳医学院多媒体形态学实验室

出　　处：《中国公共卫生》2009年第7期827-828,共2页Chinese Journal of Public Health

基　　金：国家自然科学基金(30760227);贵州省科技攻关项目[黔科合NY字(2008)3060]

摘　　要：目的扩增、克隆和表达亚洲牛带绦虫(Taenia saginata asiatica)Spef1-Like基因全长cDNA,并进行表达产物的纯化。方法以亚洲牛带绦虫成虫cDNA文库中含Spef1-Like基因的质粒作为模板,扩增该基因,将其克隆到原核表达载体pET-28a(+)中,测序鉴定重组质粒,在大肠埃希菌BL-21/DE3中用异丙基硫代-β-D半乳糖苷(IPTG)诱导,表达产物通过十二烷基磺酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)鉴定,重组后的蛋白用His-镍蛋白纯化柱纯化。结果PCR,双酶切及DNA测序见过均表明重组质粒pET-28a(+)-Spef1-Like构建成功,该基因在大肠埃希菌中以上清表达,纯化后蛋白通过SDS-PAGE鉴定结果表明该基因在大肠埃希菌BL-21/DE3得到高效表达。结论成功克隆了亚洲牛带绦虫Spef1-Like基因,并表达和纯化了Spef1-Like蛋白,为研究Spef1-Like的功能及其疫苗等研究提供了基础依据。Objective .To clone and express Taenia saginata asiatica＇ s Spefl-like gene, and purify its protein. Methods The homologous sequence of adult Taenia saginata asiatica＇ s Spefl-like gene and its full-length coding region were obtained from the full-length eDNA library by PCR and sequenced. Meanwhile the encoding sequence was cloned into the prokaryotic expression vector pET-28a（ ＋ ）. Then it was digested with endonuclease and then expressed in E. coli BL21 with IPTG induction. The recombinant product was purified according to the protocol NTA agarose and detected by SDS-PAGE. Results Spefl-like gene cDNA sequence was obtained and then successfully cloned into pET-28a（ ＋ ）vector. SDS-PAGE results showed that the gene expression took place in Escherichia coli BL21/DE3, and highly pure protein was achieved. Conclusion Taenia saginata asiatica＇ s Spell-like gene was successfully cloned and its expression and fusion protein was purified, which provides favorite conditions for further studies of the gene.

关 键 词：亚洲牛带绦虫 Spef1-Like基因 基因克隆 原核表达 

分 类 号：R383.32[医药卫生—医学寄生虫学]