Improved recovery of rabies virus from cloned cDNA using a vaccinia virus-free reverse genetics system.
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.
Ito
Naoto
N
Laboratory of Zoonotic Diseases, Division of Veterinary Medicine, Faculty of Agriculture, Gifu University, Gifu, Gifu 501-1193, Japan.
Takayama-Ito
Mutsuyo
M
Yamada
Kentaro
K
Hosokawa
Junji
J
Sugiyama
Makoto
M
Minamoto
Nobuyuki
N
D002460:Cell Line; D003001:Cloning, Molecular / Q000379:methods; D018076:DNA, Complementary / Q000235:genetics* / Q000302:isolation & purification; D015967:Gene Expression Regulation, Viral; D016679:Genome, Viral; D010957:Plasmids; D011818:Rabies / Q000821:virology*; D011820:Rabies virus / Q000235:genetics / Q000302:isolation & purification* / Q000472:pathogenicity; D011995:Recombination, Genetic
D018076:DNA, Complementary