鸡马立克氏病毒UL36^(USP)的pTYB1表达载体的构建及表达分析  

Construction and Expression of Recombinant Plasmid p TYB1 Harboring UL36^(USP) Fragment of MDV Encoding UL36 Gene

作　　者：金雯[1] 王梦云[2] 余致君 郑海南[2] 邓晨[2] 艾永兴[2] 

机构地区：[1]吉林大学动物医学学院,长春130062 [2]吉林大学动物科学学院,长春130062

出　　处：《吉林农业大学学报》2016年第2期223-227,共5页Journal of Jilin Agricultural University

基　　金：国家自然科学基金项目(31272528);教育部新世纪优秀人才支持计划项目(NCET-12-0232);教育部留学回国人员科研启动基金项目(教外司留2013-693号)

摘　　要：UL36USP是马立克氏病毒UL36基因编码的蛋白UL36上具有去泛素化酶活性的N-端部分片段(UL36USP),为获取无冗余氨基酸残基的纯净UL36USP蛋白。试验采用p TYB1表达载体与目的基因UL36USP构建重组表达质粒,并诱导融合蛋白表达,在含有还原剂DTT的缓冲液中利用Intein发挥内含肽的剪切活性,将UL36USP从融合蛋白上分离出来,实现目的蛋白UL36USP的单柱纯化,纯化后的蛋白不带有冗余的氨基酸残基,并利用UL36和Intein的特异性抗体进行了Western Blot鉴定。该研究尝试了一种能有利于难表达蛋白的可溶性表达的方法,并利用Intein标签的剪切活性分离产生纯净的目的蛋白,如一些难表达的病毒蛋白,为抗病毒和抗肿瘤疫苗的研究提供材料。UL36USP, N-terminal fragment of UL36 protein encoded by MDV, is of deubiquitination activity. The purpose of this study is to obtain purified UL36UsP without extra amino acid residue. A recombinant plasmid was constructed by cloning UL36USP gene into pTYB1 expression vector, and the expression of UL36USP-Intein fusion protein was induced. UL36USP protein without extra amino residue was obtained post specific self-cleavage of UL36USP-Intein fusion containing dithiothreitol （DTF）. Western Blot was carried out to identify two primary antibodies, respectively, against protein UL36USP and intein. the protein eluted from column was UL36USP. This study provides a method protein in reducing buffer the purified protein using The results suggested that for the expression of some protein which may be difficult to express, such as UL36USP encoded by MDV, and also prepares ma- terials for the research on vaccine against virus and carcinoma.

关 键 词：鸡马立克氏病毒 内涵肽 P TYB1 UL36 去泛素化酶 病毒蛋白 

分 类 号：S852.65[农业科学—基础兽医学]