日本血吸虫童虫文库免疫筛选及其高丰度表达膜蛋白初步鉴定

目的 获得新的可能具有诊断意义的血吸虫蛋白分子并对其鉴定。 方法 免疫学方法筛选日本血吸虫童虫cDNA文库，对获得的阳性克隆进行测序分析，设计引物体外扩增目的蛋白基因，将其亚克隆入原核表达载体pET28a中，并转化大肠埃希菌BL21，进行诱导表达，最后用Western blotting鉴定。 结果 获得34个阳性克隆，选择其中24个进行测序，其中13个是日本血吸虫相对分子质量（Mr）为22 600膜相关蛋白（Sj22.6）基因。Sj22.6基因在体外被成功扩增，并被亚克隆入pET28a中进行表达。Western Blotting 结果显示，菌体表达的条带可以被感染兔血清以及血吸虫患者血清识别。 结论 Sj22.6膜相关蛋白基因在童虫cDNA文库中可以被高频率筛出，融合蛋白Sj22.6成功地在BL21中表达并具有免疫反应性。

Immunoscreening of Schistosomulum cDNA Library of Schistosomajaponicum and Preliminary Identification of Membrane-AssociatedProtein Abundant in Tegument

Objective To obtain genes encoding the novel molecules for diagnosis of schistosomiasis.Methods Juvenile S.japonicum cDNA library was immunoscreened to obtain positive clones.By DNA sequencing and sequence analysis,the target gene was amplified by PCR and subcloned into prokaryotic plasmid pET28a.The recombinant plasmid was transformed into E.coli BL21 followed by expression of the protein induced by IPTG.The protein was identified by Western blotting.Results 34 positive clones were obtained,24 of which were chosen to be sequenced,13 of which were Sj22600 gene.The protein were recognized with sera of infected rabbits and patients with acute and chronic schistosomiasis by Western blotting.Conclusion The gene coding for Sj22600 membrane protein was screened with high frequency in the cDNA library.The E.coli BL21 transformed with the recombinant plasmid can express the fusion protein,which shows immunoactivity.