Skip to main content

Part of the book series: Methods in Molecular Medicine™ ((MIMM,volume 13))

Abstract

The SFV Expression System is a DNA expression system used to produce recombinant protein in eukaryotic cells (1,2). The SFV system is based on the Semliki Forest Vnus (SFV), which has several features that provide distinct advantages for a good cDNA expression system. These are:

  1. 1.

    The SFV RNA genome has a positive polarity and thus functions directly as an mRNA. Infectious RNA molecules can be therefore obtained by transcription from a full-length cDNA copy of the genome,

  2. 2.

    The SFV RNA molecule codes for its own RNA rephcase. Consequently, within a few hours after infection, up to 200,000 copies of the plus-RNAs are made in a single cell (3),

  3. 3

    SFV replication occurs in the cell cytoplasm, where the virus replicase transcribes and caps the subgenomes for production of the structural proteins (4) This eliminates problems, such as mRNA splicing, limitations in transcription factors, problems with capping effctency, and mRNA transport,

  4. 4.

    The cytopathic effects in infected cells appear late during infection. Thus, there is an extensive time window of approx 4–24 h after infection during which a very high expression level of the SFV structural proteins is obtained without significant morphologrcal change of host cells, and

  5. 5.

    Under laboratory conditions, SFV infects a broad range of different cultured cells mammalian, avian, reptilian, amphibian, and insect cells.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 89.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Liljestrom P. and Garoff H. (1991) A new generation of animal cell expression vectors based on the Semliki Forest Virus replicon Bio/Technology 9, 1356–1361.

    Article  PubMed  CAS  Google Scholar 

  2. Ciccarone V, Anderson D., and Jessee J. (1994) Heterologous protein expression in mammalian cells with the SFV gene expression system Focus 16, 94–98.

    Google Scholar 

  3. Wengler G. (1980) Effects of alphaviruses on host cell macromolecular synthesis, in The Togaviruses (Schlesinger R W, ed), Academic, New York, pp 459–472

    Chapter  Google Scholar 

  4. Strauss E. G. and Strauss J. H (1986) Structure and replication of the alphavinus genome, in The Togaviridae and Flaviviridae (Schlesinger S S and Schlesinger M J., eds.), Plenum, New York, pp. 35–90.

    Chapter  Google Scholar 

  5. Olkkonen V M., Liljestròm P, Garoff H, Simons K, and Dotti C G. (1993) Expression of heterologous proteins in cultured rat hippocampal neurons using the semliki forest virus vector. J Neurosci Res 35, 445–451.

    Article  PubMed  CAS  Google Scholar 

  6. Destrooper F, Simons M, Multhaup G, Vanleuven F., Beyreuther K, and Dotti C G (1995) Production of intracellular amyloid-containing fragments in hippocampal neurons expressing human amyloid precursor protein and protection against amyloidogenesis by subtle amino acid substitutions in the rodent sequence EMBO J 14, 4932–4938

    CAS  Google Scholar 

  7. Dehoop M, Vonposer C., Lange C., Ikonen E., Hunziker W., and Dotti C. G (1995) Intracellular routing of wild-type and mutated polymeric immunoglobulin receptor in hrppocampal neurons in culture. J. Cell Biol 130, 1447–1459

    Article  CAS  Google Scholar 

  8. Paul N L., Marsh M., McKeating J A., Schulz T F, Liljestrom P, Garoff H., and Weiss R. A. (1993) Expression of HIV-1 envelope glycoproteins by Semliki Forest Virus vectors. AIDS Res Hum Retroviruses 9, 963–970

    Article  PubMed  CAS  Google Scholar 

  9. Olkkonen V. M, Dupress P., Killisch I, Lutcke Zerial M., and Simons K (1993) Molecular cloning and subcellular localization of three GTP-binding proteins of the rab subfamily J Cell Sci 106, 1249–1261

    PubMed  CAS  Google Scholar 

  10. Mikus P, Urano T, Liljestrom P, and Ny T (1993) Plasminogen-activator type 2 (PAI-2) is a spontaneously polymertsing SERPIN. Eur J Biochem 218, 1071–1082

    Article  PubMed  CAS  Google Scholar 

  11. Olkkonen V M, Liljestrom P., Garoff H, Simons K, and Dotti C G (1993) Expression of heterologous proteins in cultured rat htppocampal neurons using the semliki forest virus vector. J Neurosci Res 35, 445-51.

    Google Scholar 

  12. Zhou X, Berglund P., Rhodes G, Parker S E., Jondal M, and Liljestrom P (1994) Self-replicating Semliki Forest Virus RNA as recombinant vaccine. Vaccine 12, 1510–1514

    Article  PubMed  CAS  Google Scholar 

  13. Zhou X., Berglund P, Zhao H, Liljestróm P., and Jondal M (1995) Generation of cytotoxic and humoral immune responses by nonreplicative recombinant Semliki Forest Virus Proc Natl Acad Sci USA 92, 3009–3013

    Article  PubMed  CAS  Google Scholar 

  14. Liljestrom P, Lusa S, Huylebroeck D., and Garoff H (1991) In vitro mutagenesis of a full-length cDNA clone of Semliki Forest virus the small 6,000-molecular-weight membrane protein modulates vuus release. J Virol 65, 4107–4113

    PubMed  CAS  Google Scholar 

  15. Bradish C J, Allner K., and Maber H B (1971) The virulence of original and derived strains of Semliki Forest virus for mice, gumea pigs and rabbits. J Gen Virol 12, 141–160.

    Article  PubMed  CAS  Google Scholar 

  16. Kozak M (1989) The scanning model for translatron. an update J Cell Biol. 108, 229–241

    Article  PubMed  CAS  Google Scholar 

  17. Berglund P, Sjoberg M., Garoff H., Atkins G. J, Sheahan B. J, and Liljestrom P. (1993) Semliki Forest virus expression system production of conditionally infectious recombinant particles. Bio/Technology 11, 916–920.

    Article  PubMed  CAS  Google Scholar 

  18. Levis R., Weiss B. G., Tsiang M., Huang H., and Schlesinger S. (1986) Deletion mapping of Sindbis virus DI RNAs derived from cDNAs defines the sequences essential for replication and packaging Cell 44, 137–145

    Article  PubMed  CAS  Google Scholar 

  19. Weiss B, Nitschko H., Ghattas I., Wright R., and Schlesinger S. (1989) Evidence for specificity in the encapsidation of Sindbis virus RNAs J Virol 63, 5310–5318.

    PubMed  CAS  Google Scholar 

  20. Garry R. F. (1994) Sindbis virus-induced inhibition of protein synthesis is partially reversed by medium containing an elevated potassium concentration J Gen Viral 75, 411–415.

    Article  CAS  Google Scholar 

  21. Sanes J., Rubenstem J. L. R., and Nicolas J.-F. (1986) Use of recombinant retrovirus to study post-implantation cell lineage in mouse embryos EMBO J 5, 3133–3142.

    PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 1998 Humana Press Inc.

About this protocol

Cite this protocol

Ciccarone, V.C., Jessee, J.A., Liljeström, P. (1998). The SFV Gene Expression System. In: Reischl, U. (eds) Molecular Diagnosis of Infectious Diseases. Methods in Molecular Medicine™, vol 13. Humana Press, Totowa, NJ. https://doi.org/10.1385/0-89603-485-2:237

Download citation

  • DOI: https://doi.org/10.1385/0-89603-485-2:237

  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-0-89603-485-3

  • Online ISBN: 978-1-59259-597-6

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics