Skip to main content

Identification and Isolation of Lysogens with Induced Prophage

  • Protocol
Bacteriophages

Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 501))

Abstract

The fate of lysogens following prophage induction has assumed added significance with the finding that in many pathogens virulence genes are carried on prophages and, in some, the production and/or release of the virulence factor is under control of the phage lytic regulatory program. We outline a method for identifying and characterizing from a total lysogen population, the subpopulation in which the prophage is induced. The prophage is genetically altered so that on induction it does not go through the lytic pathway, but does express a resolvase that acts at a reporter cassette located at another site on the bacterial chromosome to irreversibly change the resistance of the bacterium from tetracycline to chloramphenicol. Thus, induced derivatives survive and are easily identified even if they make up a small fraction of the population.

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

Access this chapter

Subscribe and save

Springer+ Basic
$34.99 /Month
  • Get 10 units per month
  • Download Article/Chapter or eBook
  • 1 Unit = 1 Article or 1 Chapter
  • Cancel anytime
Subscribe now

Buy Now

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 189.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 239.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 249.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

Similar content being viewed by others

References

  1. Waldor, M. K., Friedman, D. I. & Adhya, S. L. (2005) Phages; Their role in bacterial pathogenesis and biotechnology (ASM Press, Washington, D.C.).

    Google Scholar 

  2. Tyler, J. S., Livny, J. & Friedman, D. I. (2005) in Phage: Role in Pathogenesis and Biotechnology, eds. K. W. M., Friedman, D. I. & Adhya, S. (ASM Press, Washington, D.C.).

    Google Scholar 

  3. Camilli, A. & Mekalanos, J. J. (1995) Use of recombinase gene fusions to identify Vibrio cholerae genes induced during infection. Mol. Microbiol. 18, 671–683.

    Article  CAS  PubMed  Google Scholar 

  4. Livny, J. & Friedman, D. I. (2004) Characterizing spontaneous induction of Stx encoding phages using a selectable reporter system. Mol. Microbiol. 51, 1691–1704.

    Article  CAS  PubMed  Google Scholar 

  5. Eisen, H. A., Fuerst, C. R., Siminovitch, L., Thomas, R., Lambert, L., Pereira da Silva, L. & Jacob, F. (1966) Genetics and physiology of defective lysogeny in K12 (lambda): studies of early mutants. Virology 30, 224–241.

    Article  CAS  PubMed  Google Scholar 

  6. Quandt, J. & Hynes, M. F. (1993) Versatile suicide vectors which allow direct selection for gene replacement in gram-negative bacteria. Gene 127, 15–21.

    Article  CAS  PubMed  Google Scholar 

  7. Powell, B. S., Rivas, M. P., Court, D. L., Nakamura, Y. & Turnbough, C. L., Jr. (1994) Rapid confirmation of single copy lambda prophage integration by PCR. Nucleic Acids Res 22, 5765–5766.

    Article  CAS  PubMed  Google Scholar 

  8. Grindley, N. D. (1983) Transposition of Tn3 and related transposons. Cell 32, 3–5.

    Article  CAS  PubMed  Google Scholar 

  9. Yu, D., Ellis, H. M., Lee, E. C., Jenkins, N. A., Copeland, N. G. & Court, D. L. (2000) An efficient recombination system for chromosome engineering in Escherichia coli. Proc. Natl. Acad. Sci. U S A 97, 5978–5983.

    Article  CAS  PubMed  Google Scholar 

  10. Ellis, H. M., Yu, D., DiTizio, T. & Court, D. L. (2001) High efficiency mutagenesis, repair, and engineering of chromosomal DNA using single-stranded oligonucleotides. Proc. Natl. Acad. Sci. U S A 98, 6742–6746.

    Article  CAS  PubMed  Google Scholar 

  11. Lee, S. H., Hava, D. L., Waldor, M. K. & Camilli, A. (1999) Regulation and temporal expression patterns of Vibrio cholerae virulence genes during infection. Cell 99, 625–634.

    Article  CAS  PubMed  Google Scholar 

  12. Weisberg, R. A. & Landy, A. (1983) in Lambda II, eds. Hendrix, R. W., Roberts, J. W., Stahl, F. W. & Weisberg, R. A. (Cold Spring Harbor Laboratory, Cold Spring Harbor, NY), pp. 211–250.

    Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2009 Humana Press, a part of Springer Science+Business Media, LLC

About this protocol

Cite this protocol

Livny, J., LaRock, C.N., Friedman, D.I. (2009). Identification and Isolation of Lysogens with Induced Prophage. In: Clokie, M.R., Kropinski, A.M. (eds) Bacteriophages. Methods in Molecular Biology™, vol 501. Humana Press. https://doi.org/10.1007/978-1-60327-164-6_22

Download citation

  • DOI: https://doi.org/10.1007/978-1-60327-164-6_22

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-58829-682-5

  • Online ISBN: 978-1-60327-164-6

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics