Establishment and characterization of plasmid-driven minigenome rescue systems for Nipah virus: RNA polymerase I- and T7-catalyzed generation of functional paramyxoviral RNA

Alexander Freiberg, Lhia Krista Dolores, Sven Enterlein, Ramon Flick

Research output: Contribution to journalArticle

25 Scopus citations

Abstract

In this study we report the development and optimization of two minigenome rescue systems for Nipah virus, a member of the Paramyxoviridae family. One is mediated by the T7 RNA polymerase supplied either by a constitutively expressing cell line or by transfection of expression plasmids and is thus independent from infection with a helper virus. The other approach is based on RNA polymerase I-driven transcription, a unique approach for paramyxovirus reverse genetics technology. Minigenome rescue was evaluated by reporter gene activities of (i) the two different minigenome transcription systems, (ii) genomic versus antigenomic-oriented minigenomes, (iii) different ratios of the viral protein expression plasmids, and (iv) time course experiments. The high efficiency and reliability of the established systems allowed for downscaling to 96-well plates. This served as a basis for the development of a high-throughput screening system for potential antivirals that target replication and transcription of Nipah virus without the need of high bio-containment. Using this system we were able to identify two compounds that reduced minigenome activity.

Original languageEnglish (US)
Pages (from-to)33-44
Number of pages12
JournalVirology
Volume370
Issue number1
DOIs
StatePublished - Jan 5 2008

Keywords

  • Antiviral drug screening
  • High throughput
  • Nipah virus
  • Paramyxovirus
  • RNA polymerase I-driven minigenome rescue

ASJC Scopus subject areas

  • Virology

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