牛病毒性腹泻病毒HB-bd毒株E2基因的克隆表达及免疫原性分析

Cloning and expression of the E2 gene of the HB-bd strain of bovine viral diarrhoea virus and immunogenicity analysis of the E2 protein

为了研究牛病毒性腹泻病毒E2蛋白的免疫原性,试验采用PCR技术扩增牛病毒性腹泻病毒HB-bd毒株去除跨膜区的E2基因,将获得的s E2基因克隆到p ET20b载体构建重组质粒p ET20bs E2,并转化大肠杆菌BL-21(DE3)感受态细胞及诱导表达,应用SDS-PAGE、Western-blot和Dot-ELISA对表达产物进行分析,取纯化的表达产物加等量佐剂免疫家兔,观察免疫效果。结果表明:s E2基因得到了高效表达,表达的重组蛋白分子质量约为37.0 ku,表达产物能诱导免疫家兔产生高效价抗BVDV抗体。说明表达产物s E2蛋白具有良好的免疫原性。

To study the immunogenicity of the E2 protein of bovine viral diarrhea virus （BVDV）, the polymerase chain reaction （PCR） technique was used to amplify the E2 gene removed C - terminal transmernbrane domain of the HB - bd strain of BVDV. The obtained sE2 gene was cloned into the expression vector pET20b to construct a recombinant plamid pET20b -sE2, and then transfected into E. coli BL21 （DE3）. After induced expression of pET20b - sE2 fusion expression vector by isopropyl - β - D - thiogalactoside （ IPTG）, the expressed product was ana- lyzed using SDS - PAGE and Western blot and dot - enzyme linked immunosorbent assay （ Dot - ELISA）. The rabbits were immunized with the purified recombinant proteins plus the same amount of adjuvants to observe the immune effects. The result showed that the sE2 protein was highly expressed, and the expressed recombinant protein had a molecular mass of about 37kDa. The expressed product could induce the immunized rabbits to produce high - titer antibodies against BVDV. The results indicate that the expressed recombinant protein sE2 had good immunogenicity.