| 摘要: |
| (目的)为构建猪流感病毒H1N1亚型dM2蛋白的原核表达系统，获得具有生物活性的GST-dM2重组蛋白。（方法）本研究从H1N1亚型猪流感病毒核酸中克隆出M2全长基因，采用OE-PCR技术得到缺失跨膜区的M2基因（dM2），构建重组表达质粒pGEX-6p-dM2，转化大肠杆菌BL21，经IPTG诱导表达后，进行GST亲和层析柱纯化和Western- blotting鉴定。（结果）结果表明GST-dM2融合蛋白在大肠杆菌BL21中获得较高表达，并以可溶形式存在，表达量为22.75%。纯化后的GST-dM2重组蛋白为38 kD，经Western- blotting证明具有良好的反应原性。（结论）GST-dM2重组蛋白的成功获得，为进一步研究猪流感亚单位疫苗奠定了基础。 |
| 关键词: 猪流感病毒 M2 原核表达 鉴定 |
| DOI： |
| 投稿时间：2011-11-03修订日期：2011-12-12 |
| 基金项目: |
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| The Prokaryotic Expression and Identification of dM2 Protein of H1N1 subtype Swine Influenza Virus |
| SUN Yue-hui |
| (China Institute of Veterinary Drug Control,Beijing) |
| Abstract: |
| (Objective) This experiment was conducted to utilize the dM2 prokaryotic protein expression system of H1N1 subtype swine influenza virus(SIV)，and obtain biologically active recombinant protein GST-dM2. (Method)M2 gene was amplified from H1N1 subtype SIV nucleic acid，and by using OE-PCR(Overlap Extension-PCR) technology the transmembrane region of the M2 gene (dM2) was missed from gene M2. Recombinant expression plasmid pGEX-6p-dM2 was transferred into Escherichia coli BL21， induced expression by IPTG. The expression products were purified by GST affinity chromatography and identified by Western-blotting .(Result) The results showed that GST-dM2 fusion protein was in higher expression and soluble form in E. coli BL21， with the expression level of 22.75%. Western-blotting proved that the purified GST-dM2 recombinant protein of 38 kD in size shared good reactogenicity. (Conclusion) This study should provide a foundation for further study of swine influenza subunit vaccine. |
| Key words: Swine influenza virus M2 protein Prokaryotic expression Identification |
