H3N8型马流感血凝素核酸疫苗的初步研究

发布时间：2018-06-10 04:22

本文选题：流感病毒A型 + HN亚型　；参考：《中国医药生物技术》2010年03期

【摘要】：目的研发一种具有良好免疫原性的H3N8型马流感血凝素(hemagglutinin,HA)核酸疫苗。方法根据马A型流感病毒A/Equine/Xinjiang/3/08(H3N8)的HA蛋白序列,经密码子优化后化学合成能够表达相应蛋白的基因片段,并将该片段克隆到核酸疫苗载体pJW4303中,构建带有天然信号肽的H3HA核酸疫苗,命名为H3HA/XJ3-08-wt;进一步对HA基因进行修饰,以人组织纤维蛋白酶原激活剂(human tissue plasminogen activator,tPA)取代H3HA天然信号肽,命名为H3HA/XJ3-08-tPA,或切除部分HA的基因片段使之只表达HA蛋白的胞外域,命名为H3HA/XJ3-08-dTM。用这3种重组质粒分别转染293T细胞,蛋白表达经蛋白质印迹检测得到确认。采用电转录法免疫新西兰白兔。ELISA方法检测免疫后兔血清中抗H3HA抗体滴度,血凝抑制实验(hemagglutination inhibition,HI)检测保护性抗体水平。结果 3种H3HA核酸疫苗均可在293T细胞中高效表达,并能够在新西兰白兔体内诱导产生特异性抗H3HA抗体,HI检测到不同水平的保护性抗体。其中以H3HA/XJ3-08-tPA的免疫原性最强。结论新构建的H3N8型马流感血凝素核酸疫苗具有良好的免疫原性,为此类疫苗的进一步研发奠定了基础。
[Abstract]:Objective to develop a nucleic acid vaccine for H 3N 8 hemagglutinin HA with good immunogenicity. Methods according to the HA protein sequence of Ma / Equine / Xinjiangr / 3 / 08H3N8, a codon optimized gene fragment was synthesized and cloned into the nucleic acid vaccine vector pJW4303 to construct H3HA nucleic acid vaccine with natural signal peptide. H3HA / XJ3-08-wt. the HA gene was further modified to replace the natural signal peptide of H3HA with human tissue fibrinogen activator (H3HAXJ3-08-tPA), or to express only the extracellular domain of HA protein by removing part of the HA gene fragment, named H3HAXXJ3-08-dTM. The three recombinant plasmids were transfected into 293T cells, and the protein expression was confirmed by Western blotting. The titers of anti-H3HA antibody in the serum of rabbits immunized with electrotranscription and hemagglutination inhibition assay (HI) were detected by Elisa. Results all three H3HA nucleic acid vaccines were highly expressed in 293T cells and could induce the production of specific anti-H3HA antibody HI in New Zealand white rabbits. The immunogenicity of H3HA / XJ3-08-tPA was the strongest. Conclusion the newly constructed H _ 3N _ 8 hemagglutinin nucleic acid vaccine has good immunogenicity, which lays a foundation for further research and development of this vaccine.
【作者单位】：南京医科大学第一附属医院感染病科;江苏省人民医院中美疫苗中心;中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室大动物病研究室;美国麻省大学医学院内科学系;
【基金】：黑龙江省自然科学基金重点项目(ZD200812) 兽医生物技术国家重点实验室基本科研业务费项目(NKLVBP200815) 江苏省自然科学基金(BK2006728)
【分类号】：R392

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