绵羊脑多头蚴病45m基因在多头绦虫和细粒棘球绦虫不同发育阶段同源性的比较

Comparison of the homology of the 45m family genes in different developmental stages of Taenia multiceps and Echinococcus granulosus

目的研究绵羊多头蚴病45 M重组蛋白的同源性。方法将多头蚴的45 m基因片段亚克隆到pET41b表达质粒,构建45 m-pET41b原核表达系统,诱导表达45 M重组蛋白,制备小鼠高免血清,进行免疫印迹试验(Western blot);运用RT-PCR的方法,从多头绦虫的成虫cDNA中分离到45 mA。结果45 M蛋白的高免血清可被细粒棘球绦虫不同发育阶段的天然抗原所识别,均在63kD处发生交叉反应;从多头绦虫的成虫cDNA中分离到45 m基因的同源基因,命名为45 mA(Gen-Bank号为EU326106),45 m和45 mA的核酸序列和蛋白序列分别有86%和76%的同源性;45 M和45 MA蛋白与细粒棘球蚴的保护性蛋白Eg95有57%的氨基酸同源性。结论提示45 m抗原分子在多头绦虫和细粒棘球绦虫的不同发育阶段均存在,为45 M蛋白的免疫原性的分析提供了宝贵的资料。

A gene termed as 45m was isolated from the larval stage of Taenia multiceps which was similar to the 45W gene from Taenia ovis as demonstrated by PCR. This gene was successfully expressed in E. coli through the prokaryotic expression vector pET41b. The purified 45M recombinant protein was used to immunize mice. And the antiserum produced could react with the protein appearing as a 63 kDa protein band in Western blot assay, which could be isolated from cyst fluid, protoscoleces, cyst membrane and adult worms of adult Echinococcus granulosus. In addition, a gene was also isolated from adult Echinococcus multicep and this gene had 86% and 76 0% homologous similarities to the larval nucleotide and protein sequences respectively. Also, it had 57% homologous similarity in amino acid to the protective protein EG95 of E. granolosus. The antiserum could also recognize a protein band in Western blot assay isolated from different developmental stages of E. granolosus. These results suggest that the 45m antigen exists in different stages of E. rnulticep and E. granolosus and the 45M has the same epitopes in proteins of E. granulosus.