Yanagida N, Ogawa R, Li Y, Lee L F, Nazerian K
Avian Disease and Oncology Laboratory, U.S. Department of Agriculture, East Lansing, Michigan 48823.
J Virol. 1992 Mar;66(3):1402-8. doi: 10.1128/JVI.66.3.1402-1408.1992.
Two Marek's disease virus (MDV) genes, one homologous to the glycoprotein B gene of herpes simplex virus and encoding the B antigen complex and the other encoding a 38-kDa phosphorylated protein (pp38), were inserted into the fowlpox virus (FPV) genome under the control of poxvirus promoters. Randomly selected nonessential regions of FPV were used for insertion, and the vaccinia virus 7.5 kDa polypeptide gene promoter or a poxvirus synthetic promoter was used for expression of MDV genes. Gene expression in cells infected with these recombinants was highly influenced by the promoter (the synthetic promoter being more effective) but was only slightly influenced by the insertion site and by the transcription direction of the insert relative to the direction of the flanking FPV sequences. Cells infected with an FPV recombinant expressing the MDV gB gene reacted positively with a monoclonal antibody specific to this glycoprotein in an immunofluorescence assay. Immunoprecipitation of infected cell lysates showed three glycoproteins identical to those associated with the B antigen complex of MDV (100, 60, and 49 kDa). Cells infected with a recombinant expressing the pp38 gene reacted positively with an anti-pp38 monoclonal antibody in an immunofluorescence assay. The generated protein was phosphorylated and had a molecular weight similar to that of the native pp38 protein. Sera from chickens immunized with an FPV recombinant expressing the MDV glycoprotein B gene reacted with MDV-infected cells.
将两个马立克氏病病毒(MDV)基因,一个与单纯疱疹病毒糖蛋白B基因同源并编码B抗原复合物,另一个编码38 kDa磷酸化蛋白(pp38),在痘病毒启动子的控制下插入禽痘病毒(FPV)基因组。使用FPV随机选择的非必需区域进行插入,并使用痘苗病毒7.5 kDa多肽基因启动子或痘病毒合成启动子来表达MDV基因。这些重组体感染的细胞中的基因表达受启动子高度影响(合成启动子更有效),但仅受插入位点以及插入片段相对于侧翼FPV序列方向的转录方向的轻微影响。用表达MDV gB基因的FPV重组体感染的细胞在免疫荧光试验中与针对该糖蛋白的单克隆抗体发生阳性反应。感染细胞裂解物的免疫沉淀显示三种糖蛋白与那些与MDV的B抗原复合物相关的糖蛋白相同(100、60和49 kDa)。用表达pp38基因的重组体感染的细胞在免疫荧光试验中与抗pp38单克隆抗体发生阳性反应。产生的蛋白被磷酸化,并且分子量与天然pp38蛋白相似。用表达MDV糖蛋白B基因的FPV重组体免疫的鸡的血清与MDV感染的细胞发生反应。