科研动态

为分析蓝舌病I型病毒（BTV-1）VP5蛋白的免疫原性，本研究构建了载体PET-28a-VP5并转化BL21（DE3），然后进行IPTG诱导表达，以及SDS-PAGE和Western-blot分析。结果显示，重组VP5蛋白相对分子质量约为62 kDa，与预期结果一致。进一步优化条件，发现IPTG为0.2 mmol/L、温度为25℃、诱导6 h时，重组蛋白在上清中表达量最高，通过Ni柱纯化获得的重组蛋白纯度约为75%。将纯化后的重组蛋白VP5分别以40μg/只和20 μg/只免疫Balb/c小鼠，同时设置PBS为阴性对照，对三免后血清进行间接ELISA检测，发现高、低剂量免疫小鼠均能产生针对VP5蛋白的特异性抗体。用灭活BTV-1进行DOT-ELISA检测，发现其能与VP5蛋白免疫的小鼠血清发生特异性反应，说明利用大肠杆菌表达的重组蛋白VP5具有良好的免疫特性，这为进一步开展BTV相关研究奠定了基础。

Expression，Purification and Immunogenicity Analysis onVP5 Protein

of Bluetongue Virus Serotype I

In order to analyze the immunogenicity of VP5 protein of bluetonguevirus serotype I（BTV-1），in this paper，the prokaryotic expression vector PET-28a-VP5 was constructed andtransformed into BL21（DE3）.After inducible expression by IPTG，the VP5 protein waspurified and analyzed by means of SDS-PAGE and Western-blot. The results showedthat the relative molecular mass of recombinant VP5 protein was about 62 kDa，which was consistent with the expected result. It was found that，by further optimizing，the expression levelof recombinant protein in supernatant was maximum when IPTG was 0.2 mmol/L，the bacteria solution was inducted for 6 hours at the temperature of25℃，and the purity of recombinant VP5 protein throughNi-sepharose purification was about 75%. Then the purified VP5 protein wasimmunized to Balb/c mice at the doses of 40 and 20 μgper mouse respectively，meantime，PBS was set as the negative control. The serum samples were testedby indirect ELISA after being immunized for 3 times. It was concluded that thespecific antibody against VP5 protein appeared in immunized mice with high orlow dose. In addition，it was found that inactivatedBTV-1 could specifically react with the serum of immunized mice with VP5protein by DOT-ELISA test，indicating that theimmunogenicity of recombinant VP5 protein was good，whichlaid the foundation for further research on BTV.

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