Ito Naoto, Takayama-Ito Mutsuyo, Yamada Kentaro, Hosokawa Junji, Sugiyama Makoto, Minamoto Nobuyuki
Laboratory of Zoonotic Diseases, Division of Veterinary Medicine, Faculty of Agriculture, Gifu University, Gifu, Gifu 501-1193, Japan.
Microbiol Immunol. 2003;47(8):613-7. doi: 10.1111/j.1348-0421.2003.tb03424.x.
To improve efficiency of recovery of rabies virus from cloned cDNA, we established a BHK cell clone that stably expresses T7 RNA polymerase, which we named BHK/T7-9. We also constructed new helper plasmids for expression of nucleoprotein and RNA polymerase of the RC-HL strain using the pTM1 plasmid vector, which makes the T7 RNA polymerase-transcripts from the plasmid cap-independent for translation. After co-transfection of these helper plasmids and the previously constructed full-length genome plasmid of the RC-HL strain to BHK/T7-9 cells, recombinant rabies virus was efficiently recovered from the cloned cDNA.
为提高从克隆的 cDNA 中拯救狂犬病病毒的效率，我们建立了一个稳定表达 T7 RNA 聚合酶的 BHK 细胞克隆，命名为 BHK/T7-9。我们还使用 pTM1 质粒载体构建了新的辅助质粒，用于表达 RC-HL 株的核蛋白和 RNA 聚合酶，这使得质粒上的 T7 RNA 聚合酶转录本能够不依赖帽子结构进行翻译。将这些辅助质粒与先前构建的 RC-HL 株全长基因组质粒共转染至 BHK/T7-9 细胞后，从克隆的 cDNA 中高效拯救出了重组狂犬病病毒。
