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A型流感病毒PB1-F2蛋白的原核表达、纯化及鉴定

Prokaryotic Expression, Purification and Identification of the PB1-F2 Protein of Influenza A Virus
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摘要 对A型流感病毒PB1-F2蛋白进行了原核表达及纯化,获得了谷胱甘肽硫转移酶(GST)-PB1-F2融合蛋白,以用于研究流感病毒PB1-F2蛋白的功能及分子作用机制。以RT-PCR方法扩增PB1-F2基因,将其克隆至原核表达载体pGEX-6P-1上。重组质粒pGEX-6P-1-PB1-F2转化大肠杆菌(Es-cherichia coli)菌株BL21,并诱导表达,利用谷胱甘肽-Sepharose 4B亲和柱进行纯化,所得产物进行SDS-PAGE、Western blot试验鉴定。结果显示,大肠杆菌细胞经诱导,表达出了相对分子量约为36kDa的蛋白,与GST-PB1-F2融合蛋白大小相符;Western blot鉴定显示,该融合蛋白能够被抗GST的抗体特异性识别。试验在大肠杆菌表达系统中成功表达纯化了GST-PB1-F2融合蛋白,为深入研究PB1-F2蛋白的功能奠定了基础。 The prokaryotic expression and purification of the Influenza A virus PB1-F2 protein was done and the recombinant GST-PB1-F2 fusion protein was obtained in order to elucidate the function and molecular mechanism of PB1-F2 of Influenza A virus. The cDNA of PB1-F2 was obtained using RT-PCR method and cloned to prokaryotic expression vector pGEX-6P-1. The recombinant plasmid pGEX-6P-1-PB1-F2 was expressed in Escherichia coli BL21 and the products were purified by Glutathione-Sepharose 4B. The expression product was identified by SDS-PAGE and Western blot assay. The results showed that the obtained protein with relative molecular weight of 36kDa was similar to GST-PB1-F2 fusion protein according to SDS-PAGE. The fusion protein could be recognized by anti-GST antibody. The GST-PB1-F2 fusion protein was successfully expressed and purified, which laid a foundation of further study on the function of PB1-F2.
作者 李俊杰 侯佩莉 关振宏
机构地区 吉林农业大学动物科学技术学院 吉林大学人兽共患病研究所/人兽共患病教育部重点实验室
出处 《湖北农业科学》 北大核心 2010年第4期788-790,797,共4页 Hubei Agricultural Sciences
基金 国家自然科学基金项目(30800824) 吉林大学农学部引进人才启动基金(4305050102B9)
关键词 流感病毒 谷胱甘肽硫转移酶-A型流感病毒PB1-F2蛋白融合蛋白 原核表达 纯化 Influenza virus GST-PB1-F2 prokaryotic expression purification
分类号 S852.659.5 [农业科学—基础兽医学] Q785 [生物学—分子生物学]
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参考文献10

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二级参考文献12

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湖北农业科学
2010年 第4期

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