细粒棘球绦虫烯醇酶基因克隆 表达及免疫诊断研究  

Cloning,expression and immunodiagnostic evaluation of enolase from Echinococcus granulosus

作　　者：王莹[1] 张璟[1] 袁忠英[1] 周何军[1] 沈玉娟[1] 徐馀信[1] 王燕娟[1] 伍卫平[1] 曹建平[1] 

机构地区：[1]中国疾病预防控制中心寄生虫病预防控制所,卫生部寄生虫病原与媒介生物学重点实验室,世界卫生组织疟疾、血吸虫病和丝虫病合作中心,上海200025

出　　处：《中国血吸虫病防治杂志》2012年第5期549-552,556,共5页Chinese Journal of Schistosomiasis Control

基　　金：国家重大科技专项(2012ZX10004-201;2009ZX10004-201;2008ZX10004-002);国家高技术研究发展计划(863计划)(2006AA02Z444)

摘　　要：目的克隆表达细粒棘球绦虫烯醇酶(EgEno)基因,并对其免疫诊断价值进行初步评价。方法从细粒棘球绦虫cDNA中扩增目的基因,克隆入表达载体pET28a,转化E.coliBL21(DE3),经异丙基βD硫代半乳糖苷(IPTG)诱导表达后,进行SDS PAGE和免疫印迹法鉴定;采用细粒棘球蚴病及其他几种寄生虫病患者的血清和健康人血清,通过ELISA法评价重组EgEno抗原的免疫诊断效果。结果重组质粒pET28a EgEno构建成功。SDS PAGE和免疫印迹法结果显示,重组蛋白在E.coliBL21(DE3)中获得高效表达,重组蛋白EgEno相对分子量约为50 kDa,可被细粒棘球蚴病患者血清识别。EgEno对细粒棘球蚴病患者血清的免疫诊断敏感性为81.25%。结论克隆出细粒棘球绦虫EgEno基因并在E.coliBL21(DE3)中表达,重组EgEno对细粒棘球蚴病有较好的免疫诊断价值。Objective To clone and express EgEno gene of Echinococcus granulosus,and to investigate the immunogenicity and diagnostic value of recombinant EgEno. Methods Total RNAS of E. granulosus was extracted and reversedly transcripted to cDNA. EgEno gene was amplified from cDNA and inserted into vector pET28a. The recombinant plasmid pET28a？EgEno was trans？ formed into E. coli BL21（DE3）for expression under the induction of IPTG. The expressed product was identified by SDS？PAGE and Western blotting. The purified recombinant EgEno protein was detected by ELISA with the sera of cystic echinococcosis pa？ tients,healthy persons and other patients. Results The EgEno gene was successfully amplified from cDNA of E. granulosus and a fusion protein was expressed in E. coli BL21（DE3）. The molecular weight of the expressed protein was around 50 kDa. The re？ sult of Western blotting indicated that the antigenicity of the protein was specific. The sensitivity of diagnosis by ELISA for cystic echinococcosis was 81.25%. Conclusion EgEno of E. granulosus is cloned and expressed in E. coli BL21（DE3）successfully, which might be used as a candidate antigen of immunodiagnosis for cystic echinococcosis.

关 键 词：细粒棘球绦虫 EgEno基因 克隆 表达 免疫诊断 

分 类 号：R383.33[医药卫生—医学寄生虫学]