日本血吸虫泛素活化酶E1基因的克隆、表达及鉴定

Cloning,expression and identification of Ubiquitin-activating enzyme E1 of Ubiquitin protein degradation system in Schistosoma japonicum

目的扩增并表达日本血吸虫泛素活化酶E1蛋白。方法利用PCR技术得到E1基因序列,并将其克隆入原核表达载体pET28a,转化至大肠埃希菌(E.coli)BL21株,IPTG进行诱导表达,SDS-PAGE和蛋白质印迹(Western blot-ting)分析表达产物与鉴定其免疫反应性。结果SjE1基因开放阅读框长651bp,编码217个氨基酸残基,Western blotting分析结果表明,纯化的SjE1蛋白能被疫区居民血吸虫抗体阳性者血清、急性及慢性及晚期血吸虫病患者血清识别,与健康人血清无反应。结论日本血吸虫蛋白基因SjE1在体外获得表达,其表达蛋白具有一定的抗原性。

To clone and express ubiquitin-activating enzyme（El） of Schistosoma japonicum, 666 bp SjE1 gene was amplified in vitro by PCR, then, the target gene was cloned into prokaryotic plasmid pET28a（＋）. After the recombinant plasmid was transformed into E. coli BL21, it was expressed after induction with IPTG. The expressed protein was identified by SDS- PAGE and Western blotting. In these ways, SjE1 reacted positively with sera of acute, advanced and chronic schistosomiasis patients but not with sera from healthy persons, these results demonstrate that SjE1 has been expressed and shows certain antigenicity.