Efficient bunyavirus rescue from cloned cDNA

AC Lowen, C Noonan, A McLees, Richard Michael Elliott

Research output: Contribution to journalArticlepeer-review

79 Citations (Scopus)

Abstract

Bunyaviruses are trisegmented, negative-sense RNA viruses. Previously, we described a rescue system to recover infectious Bunyamwera virus (genus Orthobunyavirus) entirely from cloned cDNA (Bridgen, A. and Elliott, R.M. (1996) Proc. Nat. Acad. Sci. USA 93, 1540015404) utilizing a recombinant vaccinia virus expressing bacteriophage T7 RNA polymerase to drive intracellular transcription of transfected T7 promoter-containing plasmids. Here we report efforts to improve the efficiency of the system by comparing different methods of providing T7 polymerase. We found that a BHK-derived cell line BSR-T7/5 that constitutively expresses T7 RNA polymerase supported efficient and reproducible recovery of Bunyamwera virus, routinely generating >10(7) pfu per rescue experiment. Furthermore, we show that the virus can be recovered from transfecting cells with just three plasmids that express full-length antigenome viral RNAs, greatly simplifying the procedure. We suggest that this procedure should be applicable to viruses in other genera of the family Bunyaviridae and perhaps also to arenaviruses. (C) 2004 Elsevier Inc. All rights reserved.

Original languageEnglish
Pages (from-to)493-500
Number of pages8
JournalVirology
Volume330
Issue number2
DOIs
Publication statusPublished - 20 Dec 2004

Keywords

  • bunyavirus
  • cDNA
  • Bunyaviridae
  • virus rescue
  • reverse genetics
  • NONSTRUCTURAL PROTEIN NSS
  • VIRAL INTERFERON ANTAGONIST
  • RNA-POLYMERASE
  • BUNYAMWERA VIRUS
  • REVERSE GENETICS
  • VACCINIA VIRUS
  • FEVER VIRUS
  • SYSTEM
  • EXPRESSION
  • TRANSFECTION

Fingerprint

Dive into the research topics of 'Efficient bunyavirus rescue from cloned cDNA'. Together they form a unique fingerprint.

Cite this