Development of a reverse genetics system for a human rabies virus vaccine strain employed in China.
The CTN rabies virus was isolated from a human in China in 1953 and subsequently attenuated by multiple passaging to a vaccine strain now approved by the WHO. In this study, we describe the development of a reverse genetics system for the CTN rabies virus strain. The recombinant full-length genomic cDNA was flanked by a hammerhead ribozyme (HamRz) and the hepatitis delta virus ribozyme (HdvRz) while the non-coding G-L region was replaced with a green fluorescent protein (GFP) gene. A set of helper plasmids encoding nucleoprotein (N), phosphoprotein (P), and large protein (L) were constructed and co-transfected with recombinant full-length genome plasmid into BHK-21 cells. Recombinant virus was successfully recovered from cloned cDNA under control of the CMV promoter driven by RNA polymerase II. The recombinant virus, CTN-GFP, stably expressed GFP as detected by fluorescence microscopy. A group of 1-day-old suckling mice was challenged with the CTN-GFP strain by intracerebral inoculation, resulting in 100% morbidity and GFP expression was detected in brain tissues. The recombinant virus CTN-GFP strain recovered from cloned cDNA will be useful as a viral vector to express other foreign genes.
Huang
Ying
Y
State Key Laboratory for Molecular Virology and Genetic Engineering, Institute for Viral Disease Control and Prevention, China CDC, 100 Ying Xin St. Xuan Wu District, Beijing 100052, China.
Tang
Qing
Q
Nadin-Davis
Susan A
SA
Zhang
Shoufeng
S
Hooper
Craig D
CD
Ming
Pinggang
P
Du
Jialiang
J
Tao
Xiaoyan
X
Hu
Rongliang
R
Liang
Guodong
G
D000818:Animals; D002460:Cell Line; D002681:China; D006224:Cricetinae; D005818:Genetic Engineering / Q000379:methods*; D005827:Genetics, Microbial / Q000379:methods*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D003698:Hepatitis Delta Virus / Q000201:enzymology / Q000235:genetics; D006801:Humans; D051379:Mice; D010957:Plasmids; D016337:RNA, Catalytic / Q000235:genetics / Q000378:metabolism; D011818:Rabies / Q000821:virology; D011819:Rabies Vaccines / Q000235:genetics* / Q000276:immunology; D011820:Rabies virus / Q000235:genetics* / Q000276:immunology; D016019:Survival Analysis
D016337:RNA, Catalytic; D011819:Rabies Vaccines; D049452:Green Fluorescent Proteins