Inducible vesicular stomatitis virus (VSV) L cell line for packaging of recombinant VSV.
Recently, recombinant vesicular stomatitis viruses (VSV) have been developed as high-level expression vectors which serve as effective vaccine vectors in animals. An ideal approach for VSV vector production would be the development of stable packaging cell lines that can produce vector particles without transfection step. In this report, we describe generation of an inducible cell line that expresses the VSV polymerase gene (L) under the control of the reverse tetracycline-controlled transactivator (rtTA) system as a first step to make VSV-based packaging cell lines. Integrated polymerase (L) gene was controlled by an rtetR-dependent promoter in the rtTA-producing BHK cell line. When the cell lines were cultured in the presence of tet (tetracycline) or tetracycline derivative doxycycline, the recombinant VSV and wild type VSV were replicated, whereas in the absence of tet or tetracycline derivative doxycycline, the recombinant VSV was not replicated. Viral supernatants were harvested, diluted, and monitored by plaque assay for the presence of infectious VSV. Plaques of VSV containing an additional sequence encoding the EGFP protein allowed rapid detection of infection. Our results suggest wide applications of other surrogate viruses based on VSV. The availability of stable packaging cell lines represents a step toward the use of a VSV vector delivery system that can allow scale-up production of vector-stocks for gene therapy.
Hong
Seong-Karp
SK
Department of Microbiology, College of Medicine, The Catholic University of Korea, 137-701 Seoul, Korea.
Jung
Yong-Tae
YT
Park
Seung-Won
SW
Paik
Soon-Young
SY
D000818:Animals; D002460:Cell Line; D006224:Cricetinae; D005818:Genetic Engineering / Q000379:methods*; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000235:genetics; D012324:RNA-Dependent RNA Polymerase / Q000235:genetics*; D011993:Recombinant Fusion Proteins; D011995:Recombination, Genetic; D013752:Tetracycline / Q000378:metabolism; D014721:Vesicular stomatitis Indiana virus / Q000235:genetics* / Q000302:isolation & purification / Q000502:physiology; D010948:Viral Plaque Assay; D014764:Viral Proteins / Q000235:genetics*; D019065:Virus Assembly; D014779:Virus Replication
D011993:Recombinant Fusion Proteins; D014764:Viral Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; C040530:L protein, vesicular stomatitis virus; D012324:RNA-Dependent RNA Polymerase; D013752:Tetracycline