CAJ | 학술논문

目的构建人H5N1亚型禽流感病毒A/Anhui/1/2005 M1蛋白的原核表达系统，为进一步研究M1蛋白的生物学功能和制备其诊断试剂奠定基础。方法以该病毒基因节段七cDNA为模板，PCR扩增得到M1基因片段。将该片段亚克隆至载体pQE80-L中，构建重组质粒pQE80-L/M1，转化大肠埃希菌BL21( DE3)。IPTG诱导重组蛋白表达。金属镍离子螯合层析纯化N末端携带多聚组氨酸标签的重组M1蛋白，免疫小鼠制备多克隆抗体。结果获得了重组M1蛋白，能与抗H5N1亚型流感病毒血清发生特异性结合，且其免疫后能诱导机体产生特异性抗体。结论成功获得了人H5N1亚型禽流感病毒M1蛋白在原核细胞中高效表达。
목적 구건인H5N1아형금류감병독A/Anhui/1/2005 M1단백적원핵표체계통，위진일보연구M1단백적생물학공능화제비기진단시제전정기출。방법 이해병독기인절단칠cDNA위모판，PCR확증득도M1기인편단。장해편단아극륭지재체pQE80-L중，구건중조질립pQE80-L/M1，전화대장애희균BL21( DE3)。IPTG유도중조단백표체。금속얼리자오합층석순화N말단휴대다취조안산표첨적중조M1단백，면역소서제비다극륭항체。결과 획득료중조M1단백，능여항H5N1아형류감병독혈청발생특이성결합，차기면역후능유도궤체산생특이성항체。결론 성공획득료인H5N1아형금류감병독M1단백재원핵세포중고효표체。
Objective To generate the Escherichia coli vector expressing human H5N1 influenza virus M1 protein. To provide useful tools for detection of human H5N1 influenza virus and study on biological function of M1 protein. Methods M1 gene fragment was amplified by PCR using the influenza virus gene segment 7 as template, and was subcloned into pQE80-L vector. The recombinant plasmid pQE80-L/M1 was transformed into Escherichia coli BL21 ( DE3 ) strain. The expression of M1 was induced by isopropy-β-Dthiogalactopyranoside. We purified the recombinant M1 protein with polyhistidine tag with Ni2+ affinity chromatography. Mouse were immunized with the purified M1 protein for preparing antibodies against M1. Results The recombinant M1 protein was recognized by antiserum against H5N1 subtype influenza virus, elicit specific antibody in immunized animals. ConclusionThese confirmed that we successfully constructed the Escherichia coli vector expressing human H5N1 influenza virus M1 protein.