传染性法氏囊病毒免疫原基因在毕赤酵母中的高效分泌表达  

High Performance Secretion and Expression of vvIBDV VP2 in Pichia pastoris

作　　者：蓝胜芝 徐素珍 李宝臣 曹玉飞 赵艳敏 金顺福 

机构地区：[1]浙江诺倍威生物技术有限公司,浙江杭州310018

出　　处：《中国动物检疫》2013年第9期50-55,共6页China Animal Health Inspection

摘　　要：从国内分离的鸡传染性法氏囊病毒(IBDV)众多流行株中获得超强毒株(vvIBDV),应用RT-PCR方法,扩增得到IBDV主要保护性抗原VP2基因的开放编码框,克隆至巴斯德毕赤酵母表达载体pPICZαA,将该重组质粒pPICZαA-VP2以电转化方法转入毕赤酵母X-33感受态细胞中,通过PCR鉴定、表型筛选和抗生素浓度梯度筛选,获得高拷贝阳性重组酵母菌株X-33-pPICZαA-VP2。经摇瓶诱导表达试验获得稳定高效分泌表达的酵母工程菌,表达量为0.459mg/ml,聚丙烯酰胺凝胶电泳(SDS-PAGE)、免疫印迹(Western-blot)以及动物试验表明,重组酵母工程菌表达的目的蛋白具有鸡传染性法氏囊病病毒天然蛋白的生物活性和免疫原性。The VP2 gene of vvIBDV was inserted into the Pichia pastoris expression vector of pPICZαA. Then the recombinant plasmid of pPICZαA VP2 was transformed into X-33 by electroporation. The multi-copy insertion transformants X-33-pPICZαA-VP2 were screened by PCR 、phenotype and Zeocin-resistance. The expressed VP2 accumulated up to about 0.459mg/mL. SDS-PAGE 、Western blot and animal experiment demonstrated that VP2 product has the similar bioactivity and antigenicity as the natural one.

关 键 词：鸡传染性法氏囊病(IBD) 超强毒株VP2 巴斯德毕赤酵母 高效分泌表达 

分 类 号：S852.659.4[农业科学—基础兽医学]