鹅细小病毒VP3蛋白抗血清的制备及在鸭胚和鸭胚成纤维细胞中的增殖研究

摘要

目的 制备鹅细小病毒(GPV) VP3衣壳蛋白的兔抗血清,在绍兴麻鸭鸭胚及其成纤维细胞中成功增殖GPV.方法 根据GPVH分离株的VP3基因序列构建原核表达载体pET3a-VP3,诱导纯化VP3蛋白,皮下注射免疫新西兰白兔,收集和纯化VP3抗血清,经Western blotting检测VP3抗血清与VP3蛋白的反应性.GPV H株鹅胚尿囊液接种SPF绍兴麻鸭胚和其成纤维细胞,通过PCR反应和间接免疫荧光方法,检测GPV在鸭胚及其成纤维细胞中的增殖.结果 成功制备了VP3的兔抗血清,同时GPV在鸭胚及其成纤维细胞中进行良好增殖.结论 VP3抗血清的制备及GPV在绍兴麻鸭鸭胚及成纤维细胞中的增殖实验为GPV分子生物学特性及致病机制的研究奠定基础.%Objective To prepare antiserum against VP3 protein and propagate goose parvovims (GPV) H in duck embryo and duck embryo fibroblasts (DEFs).Methods The prokaryotic expression vector pET30a-VP3 were constructed based on VP3 seqeucne of GPV H.VP3 protein was induced,expressd and purified,then immunized the New Zealand white rabbit.The VP3 antiserum was collected,purified and verified by Westem blotting.GPV H fluid was inoculated into shaoxing duck embryo and DEFs,and the propagation of GPV was identified by PCR and indirect immunofuorescence.Result The VP3 antiserum was successfully prepared,and GPV H was well propagated in shaoxing duck embryo and DEFs.Conclusion VP3 antiserum preparation and GPV replication in duck embryo and DEFs may be provided basis for GPV molecular biological characteristic and pathogenic mechanism research.