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In Vitro Screen of Bioinformatically Selected Bacillus anthracis Vaccine Candidates by Coupled Transcription, Translation, and Immunoprecipitation Analysis

  • Orit Gat
  • Haim Grosfeld
  • Avigdor Shafferman
Part of the Methods in Molecular Biology™ book series (MIMB, volume 375)

Summary

The availability of the Bacillus anthracis genome sequence allowed for in silico selection of a few hundred open reading frames (ORFs) as putative vaccine candidates. To screen such a vast number of candidate ORFs, without resorting to laborious cloning and protein purification procedures, methods were developed for generation of PCR elements, compatible with in vitro transcription-translation and immunoprecipitation, as well as with their evaluation as DNA vaccines. Protocols will be provided for application of these methods to analyze the anti-B. anthracis antibody repertoire of hyperimmune sera or sera from convalescent and from DNA-vaccinated animals.

Key Words

In vitro transcription in vitro translation reverse vaccinology vaccine development Bacillus anthracis bioinformatics geomics DNA vaccine gene gun immunoprecipitation serological analysis immunogenicity screen antigenicity screen 

References

  1. 1.
    Mock, M. and Fouet, A. (2001) Anthrax. Annu. Rev. Microbiol. 55, 647–671.CrossRefPubMedGoogle Scholar
  2. 2.
    Friedlander, A. M., Welkos, S. L., and Ivins, B. E. (2002) Anthrax vaccines. Curr. Top. Microbiol. Immunol. 271, 33–60.PubMedGoogle Scholar
  3. 3.
    Adu-Bobie, J., Capecchi, B., Serruto, D., Rappuoli, R., and Pizza, M. (2002) Two years into reverse vaccinology. Vaccine 21, 605–610.CrossRefGoogle Scholar
  4. 4.
    Read, T. D., Peterson, S. N., Tourasse, N., et al. (2003) The genome sequence of Bacillus anthracis Ames and comparison to closely related bacteria. Nature 423, 81–86.CrossRefPubMedGoogle Scholar
  5. 5.
    Okinaka, R. T., Cloud, K., Hampton, O., et al. (1999) Sequence and organization of pXO1, the large Bacillus anthracis plasmid harboring the anthrax toxin genes. J. Bacteriol. 181, 6509–6515.PubMedGoogle Scholar
  6. 6.
    Ariel, N., Zvi, A., Makarova, K. S., et al. (2003) Genome-based bioinformatic selection of chromosomal Bacillus anthracis putative vaccine candidates coupled with proteomic identification of surface-associated antigens. Infect. Immun. 71, 4563–4579.CrossRefPubMedGoogle Scholar
  7. 7.
    Ariel, N., Zvi, A., Grosfeld, H., et al. (2002) Search for potential vaccine candidate open reading frames in the Bacillus anthracis virulence plasmid pXO1: in silico and in vitro screening. Infect. Immun. 70, 6817–6827.CrossRefPubMedGoogle Scholar
  8. 8.
    Johnson, J. L. (1994) Similarity analysis of DNAs, in Methods for General and Molecular Bacteriology, (Gerhardt, P., Murray, R. G. E., Wood, W. A., and Krieg, N. R., eds.), ASM, Washington, DC, pp. 655–682.Google Scholar
  9. 9.
    Chitlaru, T., Ariel, N., Zvi, A., et al. (2004) Identification of chromosomally encoded membranal polypeptides of Bacillus anthracis by a proteomic analysis: Prevalence of proteins containing S-layer homology domains. Proteomics 4, 677–691.CrossRefPubMedGoogle Scholar
  10. 10.
    Altboum, Z., Gozes, Y., Barnea, A., Pass, A., White, M., and Kobiler, D. (2002) Postexposure prophylaxis against anthrax: evaluation of various treatment regimens in intranasally infected guinea pigs. Infect. Immun. 70, 6231–6241.CrossRefPubMedGoogle Scholar
  11. 11.
    Grosfeld, H., Cohen, S., Bino, T., et al. (2003) Effective protective immunity to Yersinia pestis infection conferred by DNA vaccine coding for derivatives of the F1 capsular antigen. Infect. Immun. 71, 374–383.CrossRefPubMedGoogle Scholar

Copyright information

© Humana Press Inc., Totowa, NJ 2007

Authors and Affiliations

  • Orit Gat
    • 1
  • Haim Grosfeld
    • 1
  • Avigdor Shafferman
    • 1
  1. 1.Department of Biochemistry and Molecular GeneticsIsrael Institute for Biological ResearchNess-ZionaIsrael

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