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Improved Recovery of Rabies Virus from Cloned cDNA Using a Vaccinia Virus‐Free Reverse Genetics System
Author(s) -
Ito Naoto,
TakayamaIto Mutsuyo,
Yamada Kentaro,
Hosokawa Junji,
Sugiyama Makoto,
Minamoto Nobuyuki
Publication year - 2003
Publication title -
microbiology and immunology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.664
H-Index - 70
eISSN - 1348-0421
pISSN - 0385-5600
DOI - 10.1111/j.1348-0421.2003.tb03424.x
Subject(s) - t7 rna polymerase , biology , rabies virus , plasmid , virology , complementary dna , microbiology and biotechnology , virus , nucleoprotein , polymerase , vaccinia , lyssavirus , recombinant dna , rhabdoviridae , gene , genetics , bacteriophage , escherichia coli
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.