盖塔病毒非结构蛋白NSP1多克隆抗体的制备及鉴定

Preparation and identification of polyclonal antibody against nonstructural protein 1 of Getah virus

试验旨在获得能够特异性识别盖塔病毒(Getah virus, GETV)非结构蛋白NSP1的抗体,用于鉴定NSP1蛋白及其分布。采用RT-PCR技术扩增GETV的非结构蛋白NSP1基因,构建了NSP1蛋白的原核表达质粒pET-NSP1,转化至大肠杆菌BL21(DE3)诱导大量表达。纯化原核表达的目的蛋白,免疫BALB/c小鼠,制备了NSP1蛋白的多克隆抗体,间接ELISA效价达1∶10^(5)以上。Western blot和免疫荧光试验(IFA)结果显示,NSP1蛋白多克隆抗体与GETV有良好的反应性。NSP1蛋白多克隆抗体的制备,为进一步研究该蛋白在盖塔病毒复制周期中的功能奠定了基础。

The aim of the test was to obtain the antibody that could specifically recognize the nonstructural protein NSP1 of the Getah virus, and the antibody was used to identify the NSP1 protein and its distribution. The nonstructural protein(NSP1) gene of GETV was amplified by RT-PCR. The prokaryotic expression plasmid pET-NSP1 of the NSP1 protein was constructed and transformed into BL21(DE3). Then, the target protein expressed in prokaryotic expression was purified and was used to immunize BALB/c mice. And the polyclonal antibody against the NSP1 protein was prepared. The titer of the indirect ELISA was more than 1∶10^(5). Western blot and immunofluorescence assay(IFA) showed that the NSP1 protein polyclonal antibody had good reactivity with GETV. The preparation of the NSP1 polyclonal antibody laid a foundation for further study of the function of the NSP1 protein in the replication cycle of the Getah virus.