在大肠杆菌中高效表达具有抗病毒活性的可溶性犬α7干扰素  

High level expression of recombinant canine Interferon-alpha-7 with the antiviral activity in Escherichia coli

作　　者：唐博[1] 崔子寅[1] 王艺萌[1] 魏双施[1] 高明春[1] 王君伟[1] 

机构地区：[1]东北农业大学动物医学学院,黑龙江哈尔滨150030

出　　处：《中国预防兽医学报》2013年第4期321-324,共4页Chinese Journal of Preventive Veterinary Medicine

基　　金：现代农业产业技术体系建设专项资金(CARS-37)

摘　　要：为了在大肠杆菌表达系统中高效表达具有抗病毒活性的犬α干扰素(CaIFN-α),本研究从犬肝脏中PCR获得CaIFN-α第7亚型(CaIFN-α7)基因,通过重叠延伸PCR方法在CaIFN-α7的N端融合连接内含肽SspDnaB intein(SDI)基因,并将该融合基因连接至pMAL-c2X载体中,构建含有内含肽和麦芽糖结合蛋白基因的CaIFN-α重组质粒。重组质粒在大肠杆菌Rosetta中经IPTG诱导高效表达可溶性CaIFN-α蛋白,经直链淀粉树脂亲和层析纯化后可以得到纯度较高的目的蛋白。重组蛋白经MDCK-CDV检测系统证明具有生物学活性,为该干扰素的大量生产及临床应用奠定了基础。To achieve high-level expression of soluble recombinant CalFN-a7 in E.coli, a fusion gene of Ssp DnaB intein （SDI） coding sequence on the upstream of the CalFN-a7 cDNA was generated by overlap extension （SOE）-PCR, and serted into pMAL-c2X to construct the recombinant plasmid of pMAL-SDI-CalFN-a for expressing in E.coli. The SDS-PAGE analysis indicated that the expressed fusion protein was about 80 ku and mainly expressed in soluble form which was reacted positively with rabbit anti-CalFN-ct7 serum identified by western blot. Moreover, the purified fusion protein by amylose resin column possessed the antiviral activity as detecting in MDCK/CDV system.

关 键 词：犬α干扰素 内含肽 麦芽糖结合蛋白 可溶性表达 抗病毒活性 

分 类 号：S852.61[农业科学—基础兽医学]