Abstract:

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.

Key words: Echinococcus granulosus；Enolase；Clone；Expression；Immunodiagnosis

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?β?D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.coli BL21（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.coli BL21 （DE3）中表达，重组EgEno对细粒棘球蚴病有较好的免疫诊断价值。

关键词: 细粒棘球绦虫；EgEno基因；克隆；表达；免疫诊断