鸡γ-干扰素基因(ChIFN-γ)的瞬时表达及其抗病毒活性检测  

Transient Expression of Chicken Interferon Gamma Gene (ChIFN-γ) and Determination of Its Antiviral Activity

作　　者：殷喆[1,2] 张文龙[1,2] 陈伟业[2] 刘胜旺[2] 步志高[2] 王君伟[1] 吴东来[1,2] 

机构地区：[1]东北农业大学动物医学学院,哈尔滨150030 [2]中国农业科学院哈尔滨兽医研究所,国家兽医生物技术重点实验室,农业部兽医公共和卫生重点开放实验室,哈尔滨150001

出　　处：《农业生物技术学报》2009年第6期954-959,共6页Journal of Agricultural Biotechnology

基　　金：国家高科技研究发展计划(863)项目(No.2006AA10A203);黑龙江省自然科学基金重点项目(No.ZJN-0602-01)资助

摘　　要：采用PCR技术从质粒pET-ChIFN-γ中扩增得到鸡(Gallus gallus)γ-干扰素(chicken interferon gamma,ChIFN-γ)基因,将其亚克隆到真核表达载体pCAGGS中。将鉴定正确的克隆命名为pCAGGS-ChIFN-γ,体外转染鸡胚成纤维(CEF)细胞,24h后经间接免疫荧光(IFA)和免疫印迹(Western blot)检测,表达蛋白具有良好的免疫原性。然后利用表达绿色荧光蛋白(GFP)的重组水疱口炎病毒(VSV*GFP)在CEF细胞上检测表达的ChIFN-γ抗病毒活性(AVA),经检测其抗病毒活性为2×103AU/mL,并且其活性可被抗鸡IFN-γ的多克隆抗体阻断。The chicken interferon gamma （ChIFN-γ） gene was amplified from a recombinant plasmid pET-ChIFN-γ by PCR and subcloned into a eukaryotic expression vector pCAGGS and the recombinant vector was designated as pCAGGS-ChIFN-γ CEF cells were transfected by the high quality prepared pCAGGS-ChIFN-γ IFA and Western blot showed that the ChIFN-γ was expressed transiently in the CEF cells. In this study, recombinant VSV＊GFP （Vesicular stomatitis virus expressing green fluorescence protein） and CEF cells were used in the antiviral assay （AVA） of the recombinant ChIFN-γ. The recombinant ChIFN-γ could inhibit the replication of the VSV＊GFP in CEF cells and its antiviral activity were 2×10^3 AU/mL. The antiviral activity of ChIFN-γ could be inhibited by the antibodies against ChIFN-γ.

关 键 词：鸡γ-干扰素基因 瞬时表达 抗病毒活性检测 

分 类 号：S188[农业科学—农业基础科学]