含禽源启动子的禽流感病毒核蛋白表达载体的构建及其免疫原性的检测  

Construction of a Recombinant Vector Expressing Avian influenza virus Nucleoprotein with an Avian-derived Promoter and Detection of Its Immunogenicity

作　　者：王群[1] 刘光亮[1] 肖一红[1] 童铁钢[1] 白宇[1] 张维军[1] 徐树兰[1] 吴东来[1] 

机构地区：[1]农业部兽医公共卫生重点开放实验室兽医生物技术国家重点实验室中国农业科学院哈尔滨兽医研究所,哈尔滨150001

出　　处：《农业生物技术学报》2009年第1期7-11,共5页Journal of Agricultural Biotechnology

基　　金：国家高科技发展计划(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：为了研究禽流感病毒核蛋白(nucleoprotein,NP)在鸡体内所引起的免疫反应,将其NP基因从重组质粒pVAX-NP上酶切后亚克隆至含禽源启动子的真核表达载体pCAGGS中,经筛选鉴定后获得重组子pCAGGS-NP,将鉴定正确的阳性重组子体外转染真核细胞,利用间接免疫荧光法检测目的基因的瞬时表达;采用腿部肌肉注射途径将重组子免疫SPF鸡,利用ELISA检测免疫后血清抗NP蛋白抗体效价。结果表明,该重组子无论在体外还是体内均能正确表达禽流感NP蛋白,且所表达的蛋白均具有良好的免疫原性。In order to investigate the immune responses to the nucleoprotein （NP） of Avian influenza virus, NP gene cut from recombinant plasmid pVAX-NP was subcloned into a eukaryotic expression vector pCAGGS with an avian-derived promoter. The recombinant plasmid pCAGGS-NP was obtained after PCR identification and sequencing. Then the positive pCAGGS-NP was subjected to transfect eukaryocyte and to immunize SPF chickens. Indirect florescent assay （IFA） was engaged to detect the transient expression of NP, and ELISA was used to detect the serum antibody of the immunized chickens which was resistant to NP protein. The resuits showed that NP was successfully expressed both in vitro and in vivo and had good immunoreactivity.

关 键 词：禽流感 核蛋白 真核表达载体 血清抗体 

分 类 号：S188[农业科学—农业基础科学]