表达IBDV VP2融合蛋白的重组MDV的构建及其免疫特性  

Construction and Immunological Characterization of Recombinant Marek's Disease Virus Expressing IBDV VP2 Fusion Protein

作　　者：刘红梅[1] 秦爱建[1] 刘岳龙[1] 金文杰[1] 叶建强[1] 陈鸿军[1] 邵红霞[1] 李迎晓[1] 

机构地区：[1]扬州大学江苏省动物预防医学重点实验室

出　　处：《生物工程学报》2006年第3期391-396,共6页Chinese Journal of Biotechnology

基　　金：国家高技术研究与发展计划项目(No.2002AA245051);高等学校全国优秀博士学位论文作者专项基金项目(No.200256)资助~~

摘　　要：将增强型绿色荧光蛋白基因(eGFP)与鸡传染性法氏囊病病毒(IBDV)的VP2基因融合,插入马立克氏病毒(MDV)CVI988/Rispens的非必需区US10片段中,成功构建表达VP2融合蛋白的MDVCVI988转移载体pUC18-US10-VP2。将转移载体质粒与CVI988/Rispens疫苗毒共转染鸡胚成纤维细胞(CEF),筛选获得表达VP2融合蛋白的重组MDV(rMDV)。聚合酶链式反应(PCR)和间接免疫荧光实验(IFA)证明,rMDV传至第31代仍能稳定表达VP2融合蛋白。用rMDV免疫SPF鸡,进行IBDV攻毒保护试验,1日龄SPF鸡分别用1000PFU、2000PFU、5000PFU的rMDV进行免疫,33日龄用100LD50的IBDVJS超强毒进行攻毒,鸡的免疫保护率分别为50%、60%、80%。值得注意的是,5000PFU的rMDV一次免疫1日龄SPF鸡,其法氏囊组织病理损伤等级与IBD中等毒力活疫苗常规二次免疫相当(2·0/1·5),其保护效果无显著差异(p>0·05),而与非重组病毒免疫组相比较,保护效果差异显著(P<0·01),这表明构建的表达IBDVVP2融合蛋白的rMDV可以有效地为SPF鸡提供免疫保护作用。A transfer plasmid vector pUC18-US10-VP2 was first constructed by inserting the gene of the enhancer green fluorescent protein（eGFP） fused to the VP2 gene of very virulent Infectious bursal disease virus （IBDV） JS strain into the US10 fragment of the Marek＇s disease virus （MDV） CVI988/Rispens. The recombinant virus, designated as rMDV, was developed by co-transfecting CEF with the transfer plasmid vector and simultaneously infecting with the CVI988/Rispens virus. The PCR and IFA results indicated that the rMDV is stable after 31 passages. Chickens vaccinated with rMDV were protected from challenge with 100LD50 of IBDV JS. The protection ratio of the chickens vaccinated with the 1000PFU, 2000PFU, 5000PFU of the rMDV were 50%, 60%, and 80% respectively. It is interesting that the average histopathology BF lesion scores of chicken group immunized with 5000PFU of rMDV by one-time vaccination was close to that of chicken group vaccinated with IBDV live vaccine NF8 strain for twice（2.0/1.5）. There is no difference in protection between the groups（ P 〉 0.05） but significent difference between groups immunized with 5000 PFU of rMDV and with normal MDV. This demonstrated that rMDV expressing VP2 fusion protein was effective vaccine against IBDV in SPF chickens.

关 键 词：CVD88/Rispens 传染性法氏囊病病毒 重组病毒 VP2 

分 类 号：Q78[生物学—分子生物学]