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日本血吸虫Sj-Ts4样蛋白基因的克隆、表达和初步鉴定
引用本文:潘丽红,朱绍春,任翠平,王晓楠,赵志荣,刘淼,汪学龙,沈际佳.日本血吸虫Sj-Ts4样蛋白基因的克隆、表达和初步鉴定[J].安徽医科大学学报,2007,42(3):262-264.
作者姓名:潘丽红  朱绍春  任翠平  王晓楠  赵志荣  刘淼  汪学龙  沈际佳
作者单位:1. 安徽医科大学病原生物学教研室,合肥,230032;安徽省安庆卫校基础教研组,安庆,246003
2. 安徽医科大学病原生物学教研室,合肥,230032
基金项目:安徽省科技攻关计划项目(编号:06013136B);安徽省人才开发资金项目(编号:2005Z030)
摘    要:目的为了寻找血吸虫病新的免疫学诊断候选分子,在大肠杆菌中高效表达日本血吸虫Sj-Ts4样融合蛋白。方法提取日本血吸虫成虫RNA,设计引物,用RT-PCR法扩增出日本血吸虫Sj-Ts4样蛋白基因编码序列,T载体连接,将其亚克隆入PGEX-5X-1载体中,经酶切、测序证实正确后,转化入大肠杆菌DH5α并用IPTG对该重组菌诱导表达,SDS-PAGE和Western blot方法观察表达结果。结果RT-PCR法扩增出一条大小约为870bp的片段,表达质粒PGEX-5X-1-SjTs4经双酶切和以该重组质粒为模板进行PCR扩增,均可获得一条与PCR产物一致的DNA片段,诱导表达后,经SDS-PAGE可见一条约58ku大小的GST融合蛋白条带,并可与抗GST特异性抗体反应。结论本实验成功地克隆了日本血吸虫Sj-Ts4样抗原的编码基因,并在原核细胞中进行表达,为进一步验证其免疫诊断价值奠定了基础。

关 键 词:血吸虫  日本/免疫学  克隆  分子  质粒/遗传学
文章编号:1000-1492(2007)03-0262-03
修稿时间:2007-03-28

Cloning expression and identification of Schistosoma japanicum Ts4 gene
Pan Lihong, Zhu Shaochun, Ren Cuiping, et al.Cloning expression and identification of Schistosoma japanicum Ts4 gene[J].Acta Universitis Medicinalis Anhui,2007,42(3):262-264.
Authors:Pan Lihong  Zhu Shaochun  Ren Cuiping  
Institution:Dept Parasitology, Anhui Medical University, Hefei 230032
Abstract:Objective To discover the new candidate diagnosis molecular for schistosomiasis,the SjTs4 protein gene of Schistosoma japonicum were cloned and expressed in the prokaryotic cell.Methods The total RNA were extract from adult worms of S.japonicum.Primers were designed and synthesized.The cDNA encoding SjTs4 protein was amplified by RT-PCR.The product from PCR was cloned into PMD-T vector,then was subcloned into the eukaryocyte expression vector PGEX-5X-1.After induced by IPTG,the cells were collected to be analyzed by SDS-PAGE and westem blot.Results For RT-PCR,a specific band of around 870 bp was amplified.The same band was obtained by double restriction of recombinant plasmids and PCR using recombinant plasmids(PMD-SjTs4 and PGEX-SjTs4) as template.After induced by IPTG,The results of SDS-PAGE revealed that the molecular weight of fusion protein was approximately 58 kDa.Conclusion The cDNA encoding SjTs4 protein of S.japonicum is cloned and expression in the prokaryotic cell.It provides the basis for further study on value of SjTs4 immunological diagnosis.
Keywords:Schistosoma japonicum/immunology  cloning  molecular  plasmids/genetics
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