Possee R D
Virus Res. 1986 Jul;5(1):43-59. doi: 10.1016/0168-1702(86)90064-x.
A baculovirus transfer vector was constructed using a cloned copy of the Autographa californica nuclear polyhedrosis virus (AcNPV) polyhedrin gene. The ATG translation initiation codon and 32 bases of the polyhedrin mRNA 5' leader sequences were deleted and replaced with a cDNA clone of the A/PR/8/34 influenza virus haemagglutinin (HA) gene. The hybrid HA/polyhedrin gene was transferred to AcNPV and polyhedrin-negative recombinants identified which expressed HA activity. The insect cell-derived HA bound a monoclonal antibody specific for the HA gene in immunofluorescence tests and was shown to be plasma membrane-bound by haemadsorption assays. A 65 kDa protein was identified in infected cells by immunoprecipitation. The mRNA from recombinant-infected cells was analysed and revealed a major transcript of 2.95 kilobases consistent with transcription initiation and termination at the authentic polyhedrin sites. Other transcripts of different length were also detected so it was not possible to conclude which was responsible for producing the protein. The results demonstrated that AcNPV can be used to express plasma membrane-bound virus glycoproteins.
利用克隆的苜蓿银纹夜蛾核型多角体病毒（AcNPV）多角体蛋白基因构建了杆状病毒转移载体。多角体蛋白mRNA 5'前导序列的ATG翻译起始密码子和32个碱基被删除，并用A/PR/8/34流感病毒血凝素（HA）基因的cDNA克隆进行替换。将杂交的HA/多角体蛋白基因转移到AcNPV中，并鉴定出表达HA活性的多角体蛋白阴性重组体。在免疫荧光试验中，昆虫细胞来源的HA与针对HA基因的单克隆抗体结合，并通过血细胞吸附试验显示其与质膜结合。通过免疫沉淀在感染细胞中鉴定出一种65 kDa的蛋白质。对重组感染细胞的mRNA进行分析，发现一条2.95千碱基的主要转录本，与在真实多角体蛋白位点的转录起始和终止一致。还检测到其他不同长度的转录本，因此无法确定哪个转录本负责产生该蛋白质。结果表明，AcNPV可用于表达与质膜结合的病毒糖蛋白。
