表达鸡传染性法氏囊病病毒变异株VP2基因的重组嵌合型新城疫病毒的构建与鉴定  

Construction and identification of recombinant chimeric Newcastle disease virus expressing VP2 gene of infectious bursal disease virus variant strain

作　　者：乔麒龙 黄庆 王白玉 杨盼盼 赵月政 王宝玲 崔丽瑾 苗玉和 赵军[1] QIAO Qilong;HUANG Qing;WANG Baiyu;YANG Panpan;ZHAO Yuezheng;WANG Baol-ing;CUI Lijin;MIAO Yuhe;ZHAO Jun(College of Veterinary Medicine,Henan Agricultural University,Zhengzhou 450046,China;Fujian Shengivei Biotech Co.,Ltd,Nanping,Fujian 354100,China)

机构地区：[1]河南农业大学动物医学院,河南郑州450046 [2]福建圣维生物技术有限公司,福建南平354100

出　　处：《中国兽医学报》2022年第5期867-872,共6页Chinese Journal of Veterinary Science

基　　金：国家重点研发计划资助项目(2016YFD0500800)。

摘　　要：为了研发防控IBDV变异株和基因Ⅶ型NDV感染的高效廉价多联疫苗,将IBDV中国流行变异株的VP2基因共有序列插入到含有基因Ⅶ型NDV毒株F和HN基因的嵌合型NDV LaSota-ⅦF/HN全长基因组cDNA的质粒pNDFL-ⅦF/HN中,构建含有IBDV变异毒株VP2基因的重组NDV全长感染性克隆pNDFL-ⅦF/HN-VP2。将pNDFL-ⅦF/HN-VP2和辅助质粒共转染BHK-21细胞拯救重组NDV rChiLaSota-VP2。利用PCR和Western blot对重组NDV进行鉴定,并对重组病毒的生物学特性进行研究。结果显示,重组NDV rChiLaSota-VP2拯救成功并可以正确表达IBDV VP2蛋白。第25代重组NDV在鸡胚中的生物学特性研究表明,rChiLaSota-VP2保留了LaSota株在鸡胚中高滴度复制、低致病性和遗传稳定性等生物学特性。重组NDV rChiLaSota-VP2的鸡胚半数感染量(EID_(50))峰值可达10^(-8.16)/100μL,鸡胚平均致死时间(MDT)为134 h,1日龄雏鸡脑内接种致病指数(ICPI)和6周龄鸡静脉接种致病指数(IVPI)均为0。本研究构建的表达IBDV变异株VP2基因的重组NDV rChiLaSota-VP2将为研发同时防控我国流行的基因Ⅶ型NDV及IBDV变异株感染的高效廉价多联疫苗奠定基础。To develop a multivalent vaccine for preventing the infection of infectious bursal disease virus(IBDV)variant strains and genotypeⅦNDV,a recombinant infectious clone pNDFL-ⅦF/HN-VP2 was constructed by inserting the consensus sequence of VP2 gene from IBDV variant strains in China into plasmid pNDFL-ⅦF/HN containing the full length genomic cDNA of a chimeric NDV LaSota strain which the F and HN gene was replaced by the corresponding element of genotypeⅦNDV.A recombinant NDV rChiLaSota-VP2 was rescued by co-transfection of BHK-21 cell with plasmid pNDFL-ⅦF/HN-VP2 and helper plasmids.The rChiLaSota-VP2 was verified by RT-PCR and Western blot.The biological properties of the rChiLaSota-VP2 were also investigated.The results showed that the rChiLaSota-VP2 was successfully rescued and the IBDV VP2 protein was correctly expressed by the rChiLaSota-VP2.The biological properties of the 25 th passage of the rChiLaSota-VP2 in chicken embryonated eggs demonstrated that rChiLaSota-VP2 maintained the biological properties of NDV LaSota strain,such as the efficient replication ability in chicken embryonated eggs,low pathogenicity and genetic stability.The 50%embryo infectious dose(EID_(50))of rChiLaSota-VP2reached 10^(-8.16)/100μL at 96hpost inoculation.The mean death times(MDT)of chicken embryos for the rChiLaSota-VP2was 134h,the intracerebral pathogenicity index(ICPI)in one-day-old chicks and the intravenous pathogenicity index(IVPI)in 6-week-old chickens was 0.The rChiLaSota-VP2expressing VP2gene of IBDV Chinese variant strains will lay a foundation for developing more effective and cheaper multivalent vaccine against infection of IBDV variant strain and genotypeⅦNDV.

关 键 词：鸡传染性法氏囊病毒 VP2蛋白 重组嵌合型新城疫病毒 基因Ⅶ型NDV 

分 类 号：S852.65[农业科学—基础兽医学]