Omata T, Kohara M, Sakai Y, Kameda A, Imura N, Nomoto A
Gene. 1984 Dec;32(1-2):1-10. doi: 10.1016/0378-1119(84)90026-x.
A complete cDNA copy of the genome of the attenuated type 1 poliovirus vaccine (Sabin 1) strain was constructed and inserted into the EcoRI site of the plasmid pBR325. When cultured mammalian cells were transfected with this recombinant plasmid, 20 to 50 poliovirus plaques per 10 micrograms plasmid DNA were observed. Fingerprints of the RNA of the recovered virus showed no changes when compared with those of the parental virus genome, an observation indicating that the primary structure of the cloned cDNA is a reflection of authentic poliovirus RNA. The recovered virus had the same properties as those of the Sabin 1 strain in regard to antigenicity, sensitivity to temperature, and dependency on bicarbonate concentration. These results suggest that the virus obtained by DNA transfection is indistinguishable from the Sabin 1 strain. The recombinant plasmid could therefore be used as a stable repository of the virus and as inoculum for the oral polio live vaccine.
构建了减毒1型脊髓灰质炎病毒疫苗(萨宾1株)基因组的完整cDNA拷贝,并将其插入质粒pBR325的EcoRI位点。用该重组质粒转染培养的哺乳动物细胞时,每10微克质粒DNA可观察到20至50个脊髓灰质炎病毒蚀斑。回收病毒RNA的指纹图谱与亲代病毒基因组的指纹图谱相比没有变化,这一观察结果表明克隆的cDNA的一级结构反映了真实的脊髓灰质炎病毒RNA。回收的病毒在抗原性、对温度的敏感性以及对碳酸氢盐浓度的依赖性方面与萨宾1株具有相同的特性。这些结果表明,通过DNA转染获得的病毒与萨宾1株无法区分。因此,该重组质粒可作为病毒的稳定储存库和口服脊髓灰质炎活疫苗的接种物。