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

Virology. 2008 Jan 5;370(1):33-44. doi: 10.1016/j.virol.2007.08.008. Epub 2007 Sep 27.


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.

Publication types

  • Evaluation Study
  • Research Support, N.I.H., Extramural

MeSH terms

  • Antiviral Agents / pharmacology
  • Cell Line
  • Chloramphenicol O-Acetyltransferase / metabolism
  • DNA-Directed RNA Polymerases / genetics
  • DNA-Directed RNA Polymerases / metabolism*
  • Genome, Viral / genetics*
  • HeLa Cells
  • Humans
  • Microbial Sensitivity Tests / methods
  • Nipah Virus / drug effects
  • Nipah Virus / genetics
  • Nipah Virus / physiology*
  • Paramyxoviridae / genetics
  • Paramyxoviridae / metabolism
  • Plasmids / genetics*
  • RNA Polymerase I / genetics
  • RNA Polymerase I / metabolism*
  • RNA, Viral / drug effects
  • RNA, Viral / genetics
  • RNA, Viral / metabolism*
  • Transcription, Genetic / drug effects
  • Transfection
  • Viral Proteins / genetics
  • Viral Proteins / metabolism*
  • Virus Replication / drug effects


  • Antiviral Agents
  • RNA, Viral
  • Viral Proteins
  • Chloramphenicol O-Acetyltransferase
  • bacteriophage T7 RNA polymerase
  • DNA-Directed RNA Polymerases
  • RNA Polymerase I