SARS相关冠状病毒S1融合蛋白的原核表达与纯化

目的：克隆严重急性呼吸综合征相关冠状病毒(SARS-CoV)S1蛋白编码DNA，构建原核表达质粒pGEX-5T／S1，并诱导表达谷胱甘肽巯基转移酶融合蛋白。方法：采用PCR方法扩增合成S1片段，并克隆入载体中，经双酶切鉴定和测序后把S1序列定向插入原核表达载体pGEX-5T的多克隆位点，转化大肠杆菌K802，将纯化后的融合蛋白用于SARS—CoV抗体阳性血清的检测。结果：GST—S1融合蛋白以可溶形式表达，纯化后的蛋白用ELISA法检测，结果与对照相符。结论：成功诱导原核表达并纬化出融合蛋白S1，为将其应用于SARS—CoV的特异性检测和亚单位疫苗研究奠定了基础。

Eukaryotic expression and purification of SARS coronavirus protein S1

Objective: To clone protein S1 encoding DNA of SARS coronavirus (SARS CoV) and construct the expression vector pGEX 5T/S1 to express GST S1 fusion protein.Methods: The S1 fragment was amplified by PCR and was cloned into TMD 18T vector.After verified by sequencing,the recombinant was transformed into the prokaryotic expression vector pGEX 5T.The recombinant vector was transformed into E.coli K802 and the expressed fusion protein was detected by antibody positive serum. Results: The fusion protein was expressed in soluble form; the purified protein was detected by ELISA, complying with the control method. Conclusion: Expression and purification of prokaryotic GST S1 fusion protein provide a foundation to obtain a large quantity of recombinant S1 for experimental and clinical study of SARS CoV.