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目的 在大肠杆菌中融合表达含主要抗原位点的汉滩病毒囊膜糖蛋白 G1与 NP片段 .方法 将汉滩病毒 76 - 118株 M基因编码 G1的片段与 S基因编码区 5′端约 0 .7kb的片段连接 ,克隆入 p GEX- 4T2 ,构建嵌合基因原核表达载体 p GEX- 4T2 -G1S0 .7,并在大肠杆菌 XL1- Blue中 ,诱导 GST- G1S0 .7融合蛋白的表达 .表达产物用 EL ISA和 Western blot进行鉴定 .结果 成功地构建了嵌合原核表达载体 p GEX- 4T2 - G1S0 .7.经 IPTG诱导后 ,EL ISA活性测定结果表明 ,该融合蛋白可与抗汉滩病毒 NP及糖蛋白的 m Ab特异性结合 . Western blot结果显示 ,诱导出相对分子质量 (Mr)大于 1× 10 5 的 G1S0 .7与 GST的融合蛋白 .结论 获得具有特异性结合活性的融合蛋白 GST- G1S0 .7,为汉滩病毒基因工程疫苗的研制奠定了基础 .
Objective To express glycoprotein G1 and NP of Hantaan virus glycoprotein containing the major antigenic site in Escherichia coli.Methods The Hantaan virus strain 76-118 was cloned into G1 segment of M gene and the 5 ’ .7kb fragment was cloned into pGEX-4T2 to construct chimeric prokaryotic expression vector pGEX-4T2-G1S0.7, and in E. coli XL1-Blue induced GST-G1S0.7 fusion protein expression. The product was identified by EL ISA and Western blot.Results The chimeric prokaryotic expression vector pGEX-4T2 - G1S0.7 was constructed successfully.The results of ELISA assay showed that the fusion protein could bind to anti-Hantaan virus NP and glycoprotein m Ab.The results of Western blot showed that the fusion protein G1S0.7 with GST with a molecular weight of more than 1 × 10 5 was induced.Conclusion The fusion protein GST with specific binding activity - G1S0.7, laid the foundation for the development of Hantaan virus genetic engineering vaccine.