马Viperin的抗EIAV功能初步研究及其重组腺病毒的构建  

The inhibitory activity of eViperin to EIAV and construction of recombinant adenovirus expressing the recombinant protein

作　　者：朱春辉[1] 汤艳东[2] 那雷[2] 付丽华[2] 王雪峰[2] 林跃智[2] 杜承[2] 周建华[2] 徐方[1] 

机构地区：[1]宁夏医科大学基础医学院,宁夏银川750004 [2]中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室/大动物病研究室,黑龙江哈尔滨150001

出　　处：《中国预防兽医学报》2014年第5期346-349,共4页Chinese Journal of Preventive Veterinary Medicine

基　　金：国家十二五重大新药创制(2011ZX09307-303);国家自然科学基金面上项目(31070809);兽医生物技术国家重点实验基本科研业务费(SKLVBP201312)

摘　　要：Viperin是一种细胞内抗病毒蛋白,可以被I型干扰素、多聚I:C、脂多糖及多种病毒诱导表达,在细胞内主要定位在内质网和脂滴,具有广谱的抗病毒功能。为研究马Viperin(eViperin)在抗病毒感染中的作用,本研究应用RT-PCR从马巨噬细胞中扩增eViperin基因,并将其克隆于真核表达载体pDC315中构建重组质粒pDC-eViperin-Flag,将重组质粒转染293T细胞,western blot检测结果表明eViperin在293T细胞中正确表达。将该重组表达质粒与表达马传染性贫血病毒(EIAV)Gag前体蛋白(Gag precursor,Pr55gag)的表达质粒共转染293T细胞,转染48 h后western blot检测,结果显示实验组细胞培养上清中的Pr55gag含量显著低于空白对照组,表明eViperin具有抑制EIAV释放的作用。此外,将该重组表达质粒与腺病毒骨架质粒pBHGlox△E1,E3Cre共转染HEK293细胞,进一步构建拯救出表达eViperin的重组腺病毒,为研究eViperin的抗病毒功能研究奠定了基础。Viperin is an antiviral protein which is able to be induced by virus, type I interferons, poly（I：C） and lipopolysaccharide. To explore the role of equine （e）Viperin in virus infection, the eViperin gene was amplified and cloned into pDC315 to construct the recombinant eukaryotic expression plasmid of pDC-eViperin-Flag. The pDC315-viperin-Flag was co-transfected into 293T cells with the pDRVI-SV1.0-Gag-Synwhich expressed the Gag procurer Pr55gag of equine infectious anemia virus （EIAV）, and the cell-free supernatant was collected after 48 hours and detected by western blot, and the Pr55gag in the medium of cells transfected with the pDC315-viperin-Flag was significantly lower than that of cells transfected with the empty vector. The results indicated that the eViperin protein was able to significantly inhibit the release of EIAV. In addition, the eViperin-expressing recombinant adenoviruswas generated by co-transfection of pDC315-viperin-Flag with an adenovirus-based vector pBHGlox△E1, E3Cre into HEK293 cells. which could be a useful tool for further study on eViperin.

关 键 词：Viperin 马 马传染性贫血病毒 重组腺病毒 

分 类 号：S852.65[农业科学—基础兽医学]