【摘要】 利用逆转录聚合酶链式反应(RT-PCR)方法扩增得到肠道病毒71型(EV71SHZH03)外壳蛋白VP1基因,经序列测定证实后,构建重组表达质粒VP1/pPIC9K,转化Pichiapastoris酵母宿主菌GS115,以Myc-Tag多克隆抗体作为一抗,利用双层滤膜法筛选酵母转化子.甲醇诱导表达.SDS-PAGE分析显示:表达产物的分子量约为30000,与天然VP1大小一致;ELISA实验表明,表达上清液可与EV71患者急性感染期血清呈阳性反应,表明重组蛋白VP1具有免疫原性.更多还原

【Abstract】 The gene encoding the entire capsid protein VP1 of Enterovirus 71SHZH03 was amplified by RT-PCR. After confirmed by sequence analysis, VP1 was expressed in Pichia pastoris (GS115). In situ two film method was used to screen Pichia pastoris transformants expressing high level protein of VP1. SDS-PAGE showed that the gene vp1 could express a 30000 equivalent to the natural protein. ELISA analysis of the culture supernatants confirmed that the protein had the desired immunogenicity of EV71.更多还原