Login processing...

Trial ends in Request Full Access Tell Your Colleague About Jove
Click here for the English version

Immunology and Infection

Reverse genetikk Mediated Gjenvinning av Infeksiøs murin Norovirus

doi: 10.3791/4145 Published: June 24, 2012
* These authors contributed equally


Name Company Catalog Number Comments
Lithium chloride precipitation solution Ambion AM9480
RNA storage solution Ambion AM7000
ScriptCap m7G Capping System Epicentre Biotechnologies SCCE0610
Neon transfection system Invitrogen MPK5000
Neon transfection system kit Invitrogen MPK1025
Opti-MEM I Invitrogen 31985070
Lipofectamine 2000 Transfection Reagent Invitrogen 11668-027
Agilent RNA 600 Nano kit Agilent Technologies 5067-1511
Agilent 2100 bioanalyzer Agilent Technologies G2939AA



  1. Outbreaks of gastroenteritis associated with noroviruses on cruise ships--United States. MMWR Morb. Mortal. Wkly. Rep. 51, 1112-1115 (2002).
  2. Lopman, B. A. Epidemiology and cost of nosocomial gastroenteritis, Avon, England, 2002-2003. Emerg. Infect. Dis. 10, 2002-2003 (2004).
  3. Duizer, E. Laboratory efforts to cultivate noroviruses. J. Gen. Virol. 85, 79-87 (2004).
  4. Karst, S. M., Wobus, C. E., Lay, M., Davidson, J., Virgin, H. W. T. STAT1-dependent innate immunity to a Norwalk-like virus. Science. 299, 1575-1575 (2003).
  5. Wobus, C. E. Replication of Norovirus in cell culture reveals a tropism for dendritic cells and macrophages. PLoS Biol. 2, e432 (2004).
  6. Bok, K. Inhibition of norovirus replication by morpholino oligomers targeting the 5'-end of the genome. Virology. 380, 328-337 (2008).
  7. Kim, Y., Thapa, M., Hua, D. H., Chang, K. O. Biodegradable nanogels for oral delivery of interferon for norovirus infection. Antiviral Res. 89, 165-173 (2011).
  8. McFadden, N. Norovirus Regulation of the Innate Immune Response and Apoptosis Occurs via the Product of the Alternative Open Reading Frame 4. PLoS Pathog. 7, e1002413 (2011).
  9. Kormelink, R., van Poelwijk, F., Peters, D., Goldbach, R. Non-viral heterogeneous sequences at the 5' ends of tomato spotted wilt virus mRNAs. J. Gen. Virol. 73, 2125-2128 (1992).
  10. Shuman, S., Hurwitz, J. Mechanism of mRNA capping by vaccinia virus guanylyltransferase: characterization of an enzyme--guanylate intermediate. Proc. Natl. Acad. Sci. U.S.A. 78, 187-191 (1981).
  11. Jang, S. K., Pestova, T. V., Hellen, C. U., Witherell, G. W., Wimmer, E. Cap-independent translation of picornavirus RNAs: structure and function of the internal ribosomal entry site. Enzyme. 44, 292-309 (1990).
  12. Chaudhry, Y. Caliciviruses differ in their functional requirements for eIF4F components. J. Biol. Chem. 281, 25315-25325 (2006).
  13. Goodfellow, I. Calicivirus translation initiation requires an interaction between VPg and eIF 4. E. EMBO Rep. 6, 968-972 (2005).
  14. Herbert, T. P., Brierley, I., Brown, T. D. Identification of a protein linked to the genomic and subgenomic mRNAs of feline calicivirus and its role in translation. J. Gen. Virol. 78 ( Pt5 ), 1033-1040 (1997).
  15. Chaudhry, Y., Skinner, M. A., Goodfellow, I. G. Recovery of genetically defined murine norovirus in tissue culture by using a fowlpox virus expressing T7 RNA polymerase. J. Gen. Virol. 88, 2091-20100 (2007).
  16. Ward, V. K. Recovery of infectious murine norovirus using pol II-driven expression of full-length cDNA. Proc. Natl. Acad. Sci. U.S.A. 104, 11050-11055 (2007).
  17. Yunus, M. A., Chung, L. M., Chaudhry, Y., Bailey, D., Goodfellow, I. Development of an optimized RNA-based murine norovirus reverse genetics system. J. Virol. Methods. 169, 112-118 (2010).
  18. Arias, A., Perales, C., Escarmis, C., Domingo, E. Deletion mutants of VPg reveal new cytopathology determinants in a picornavirus. PLoS One. 5, e10735 (2010).
  19. Werf, S. vander, Bradley, J., Wimmer, E., Studier, F. W., Dunn, J. J. Synthesis of infectious poliovirus RNA by purified T7 RNA polymerase. Proc. Natl. Acad. Sci. U.S.A. 83, 2330-2334 (1986).
  20. Bailey, D., Thackray, L. B., Goodfellow, I. G. A single amino acid substitution in the murine norovirus capsid protein is sufficient for attenuation in vivo. J. Virol. 82, 7725-7728 (2008).
  21. Bailey, D. Functional analysis of RNA structures present at the 3' extremity of the murine norovirus genome: the variable polypyrimidine tract plays a role in viral virulence. J. Virol. 84, 2859-2870 (2010).
Reverse genetikk Mediated Gjenvinning av Infeksiøs murin Norovirus
Play Video

Cite this Article

Arias, A., Ureña, L., Thorne, L., Yunus, M. A., Goodfellow, I. Reverse Genetics Mediated Recovery of Infectious Murine Norovirus. J. Vis. Exp. (64), e4145, doi:10.3791/4145 (2012).More

Arias, A., Ureña, L., Thorne, L., Yunus, M. A., Goodfellow, I. Reverse Genetics Mediated Recovery of Infectious Murine Norovirus. J. Vis. Exp. (64), e4145, doi:10.3791/4145 (2012).

Copy Citation Download Citation Reprints and Permissions
View Video

Get cutting-edge science videos from JoVE sent straight to your inbox every month.

Waiting X
Simple Hit Counter