猪轮状病毒重组VP8*蛋白的原核表达及免疫原性分析

Prokaryotic Expression and Immunogenic Analysis of △VP8* of Porcine Rotavirus Gottfried Strain

为了分析原核表达的猪轮状病毒（PRV）Gottfried株截短的VP8＊（△VP8＊aa64—223）重组蛋白的免疫原性，通过构建原核表达载体pET-28a—Gotffried—△VP8＊（aa64—223），转化大肠杆菌BL21（DE3）感受态细胞，IPTG诱导表达、经SDS—PAGE和Westernblotting检测，并纯化后，肌肉免疫Balb／c小鼠，证明重组蛋白相对分子质量约25．0kDa，以包涵体和可溶性两种形式存在，纯化的重组蛋白纯度在95％以上，可溶性蛋白产量可达14～15mg·L-1。ELISA方法检测表明该重组蛋白可以诱导高滴度的特异性抗体，证明其具有良好的免疫原性，为进一步研制PRV亚单位疫苗及建立ELISA诊断方法奠定基础。

The immunogenicity of truncated VP8＊ （△VP8＊,64-223 amino acids） of porcine rotavirus （PRV） Gottfried strain recombinant protein was characterized in this study by the construction of recombinant expression plasmid pET-28a-Gottfried- △VP8＊,the induction of recombinant protein with IPTG,detected by SDS-PAGE and Western blotting,and followed by immunization to Balb/c mice intramuscularly. The results demonstrated that the recombinant porcine rotavirus protein, around 25 000 Dalton, existed in both soluble and inclusion body forms and the purity of recombinant protein reached higher than 95% and the yield of soluble protein was 14-15 mg per liter. The indirect ELISA showed that the recombinant protein induced high titer of specific antibody and had good immunogenicity,which provided the potential development of PRV subunit vaccine and ELISA assay.