猪IL-2成熟蛋白在昆虫细胞中的表达及生物学活性鉴定  

Expression of porcine interleukin-2 in insect cells and identification of bioactivity of its expressed protein

作　　者：陈红英[1] 郭小参[1,2] 崔保安[1] 王彦彬[1] 张红英[1] 方剑玉 

机构地区：[1]河南农业大学牧医工程学院,河南郑州450002 [2]信阳市动物疫病预防控制中心,河南信阳464002

出　　处：《中国兽医学报》2010年第6期789-792,共4页Chinese Journal of Veterinary Science

基　　金：国家"十一五"科技支撑计划资助项目(2006BAD06A08)

摘　　要：以含猪IL-2基因的重组质粒pGEM-pIL2为模板,PCR扩增猪IL-2成熟蛋白基因。将猪IL-2成熟蛋白基因定向克隆到杆状病毒转移载体pFastBac Dual中,转化含穿梭载体Bacmid的感受态细胞DH10Bac,经抗性及蓝白斑筛选得到含猪IL-2基因的重组质粒rBacmid-IL2,转染昆虫细胞。间接免疫荧光试验表明重组猪IL-2在Sf9昆虫细胞中获得了表达,SDS-PAGE可检测到相对分子质量为20000的重组蛋白,Western-blot证实该重组蛋白可与兔抗猪IL-2单克隆抗体发生特异性反应,重组猪IL-2经纯化后能明显促进猪T淋巴细胞转化。结果表明,重组猪IL-2在昆虫细胞中得到表达,表达的重组蛋白具有一定生物学活性,为进一步开发研制新型免疫佐剂奠定了基础。Porcine IL-2 mature protein gene was amplified with recombination plasmid pGEM-pIL2 as a template,and cloned into the baculovirus transfer vector of pFastBacDual and transposed into E.coli DH10Bac which contains a shuttle vector of bacmid.The expression of the recombinant porcine IL-2 protein in Sf9 insect cells was confirmed by indirect immunofluorescence.The expressed protein was about 20 000 by SDS-PAGE and it could react to rabbit against porcine IL-2 monoclonal antibody in Western-blot.The recombinant porcine IL-2 protein induced in vitro proliferation of ConA-stimulated porcine splenocytes in MTT assay.The results showed that the recombinant porcine IL-2 protein could be expressed in Sf9 insect cells and possesses biological activities.The study paves a way of the development of new immunologic adjuvant.

关 键 词：猪白细胞介素-2 昆虫细胞 表达 生物学活性 

分 类 号：S852.23[农业科学—基础兽医学]