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目的:探讨犬细小病毒VP2亲水性编码区的免疫原性,为进一步研究基因工程亚单位疫苗奠定基础。方法:利用蛋白质分析软件Protean对已克隆的犬细小病毒VP2基因序列进行分析,选择亲水性好、抗原性强的293-520位氨基酸区域(命名为VP2S)作为靶序列,然后以已有VP2序列作为模版通过PCR扩增的方法获得VP2S,将VP2S克隆入pQE-31载体获得pQE-31-VP2S;将pQE-31-VP2S的原核表达产物经Western-blotting确认后免疫小鼠,用血凝抑制试验测定抗体水平。结果:293～520位氨基酸区域的亲水性好、抗原性强;重组质粒pQE-31-VP2S可成功表达大约29KDa的能被CPV抗血清识别的VP2S;VP2S能诱导小鼠产生高滴度的血凝抑制(HI)抗体(25)。结论:VP2S具有较强的免疫原性,能作为基因工程亚单位疫苗进行开发研究。
OBJECTIVE: To investigate the immunogenicity of the VP2 coding region of canine parvovirus (VP2) and lay a foundation for the further study of genetic engineering subunit vaccine. Methods: The VP2 gene sequence of the cloned canine parvovirus was analyzed by protein analysis software Protean. The region of amino acids 293-520 (named as VP2S) with good hydrophilicity and strong antigenicity was selected as the target sequence. Then, VP2S was cloned into pQE-31 vector to obtain pQE-31-VP2S. The prokaryotic expression product of pQE-31-VP2S was confirmed by Western-blotting and immunized mice with hemagglutination Inhibition test to determine antibody levels. Results: The amino acids 293-520 were hydrophilic and antigenic. The recombinant plasmid pQE-31-VP2S could express approximately 29KDa VP2S which can be recognized by CPV antiserum. VP2S could induce mice to produce high titer Hemagglutination inhibition (HI) antibody (25). Conclusion: VP2S is highly immunogenic and can be used as a genetic engineering subunit vaccine.