禽网状内皮组织增生病病毒env蛋白在毕赤酵母中的表达及其免疫原性研究  

The expression and immunogenicity of reticuloendotheliosis virus Env protein in Pichia pastoris

作　　者：杨彦超 王静[1] 李凯[1] 祁小乐[1] 崔红玉[1] 高玉龙[1] 刘长军[1] 高宏雷[1] 王笑梅[1] YANG Yan-chao;WANG Jing;LI Kai;QI Xiao-le;CUI Hong-yu;GAO Yu-long;LIU Chang-jun;GA0Hong-lei;WANG Xiao-mei(Division of Avian Infectious Diseases,State Key Laboratory of Veterinary Biotechnology,Harbin Veterinary Research Institute, the Chinese Academy of Agricultural Sciences,Harbin 150069,China)

机构地区：[1]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,黑龙江哈尔滨150069

出　　处：《中国预防兽医学报》2018年第12期1147-1151,共5页Chinese Journal of Preventive Veterinary Medicine

基　　金：国家肉鸡产业技术体系(CARS-42-G07)

摘　　要：为评价禽网状内皮组织增生病病毒(REV) env蛋白的免疫原性,本研究将env基因克隆至毕赤酵母表达载体pAO815,构建了含有两个env表达盒(已突变env基因中BglⅡ和Bam HⅠ酶切位点)的重组表达质粒pAO815-2env,并电转至毕赤酵母宿主菌GS115,构建重组酵母菌株pAO815-2env/GS115,经发酵培养得到的env蛋白(蛋白含量达到4.8 g/L)经亲和层析纯化后免疫SPF鸡,利用ELISA试剂盒和中和试验检测血清中的REV抗体。结果显示,重组酵母菌株表达的env蛋白,不仅能够诱导SPF鸡产生抗REV抗体,还能诱导其产生中和抗体,且产生的中和抗体效价较高,具有良好的免疫原性。本研究为开展禽网状内皮组织增生病亚单位疫苗的研制提供了实验依据。To investigate the immunogenicity of reticuloendotheliosis virus (REV)Env protein,in this study,the env gene was cloned into pAO815 vector to construct recombinant plasmid pAO815-2env containing two env expression cassettes (Bgl II and BamH I restriction sites in the env gene have been mutated).Then it was transformed into Pichia pastoris strain GSll5 by eleetroporation to construct recombinant P.pastoris strain GSll5/pAO815-2env.The expressed recombinant protein (protein content reaches 4.8g/L)was purified by affinity chromatography and then vaccinated to SPF chickens.The antibody against REV was detected by ELISA kit and neutralization assay.Results showed that the Env protein expressed by the recombinant P.pastoris strain was able to efficiently induce antibody against REV in chickens.This study provides experimental basis for the development of reticuloendotheliosis subunit vaccine.

关 键 词：禽网状内皮组织增生病病毒 ENV基因 毕赤酵母 发酵 

分 类 号：S852.65[农业科学—基础兽医学]