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Efficient allelic exchange and transposon mutagenesis in Mycobacterium tuberculosis
Open accessAbstract
A better understanding of Mycobacterium tuberculosis virulence mechanisms is highly dependent on the design of efficient mutagenesis systems. A system enabling the positive selection of insertional mutants having lost the delivery vector was developed. It uses ts- sacB vectors, which combine the counterselective properties of the sacB gene and a mycobacterial thermosensitive origin of replication and can therefore be efficiently counterselected on sucrose at 39°C. This methodology allowed the construction of M. tuberculosis transposition mutant libraries. Greater than 10 6 mutants were obtained, far exceeding the number theoretically required to obtain at least one insertion in every nonessential gene. This system is also efficient for gene exchange mutagenesis as demonstrated with the purC gene: 100% of the selected clones were allelic exchange mutants. Therefore, a single, simple methodology has enabled us to develop powerful mutagenesis systems, the lack of which was a major obstacle to the genetic characterization of M. tuberculosis .
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Citations
Cited by
Cell-Mediated Immune Responses in Tuberculosis
Cooper, A. M. (2009) · Annual Review of Immunology · doi:10.1146/annurev.immunol.021908.132703 · Cited by 1,162
Mycobacterium tuberculosis Pathogenesis and Molecular Determinants of Virulence
Smith, I. (2003) · Clinical Microbiology Reviews · doi:10.1128/cmr.16.3.463-496.2003 · Cited by 1,154
De Voss, J. J., Rutter, K., Schroeder, B. G., Su, H., Zhu, Y., & Barry, C. E. (2000) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.97.3.1252 · Cited by 532
Parish, T., & Stoker, N. G. (2000) · Microbiology · doi:10.1099/00221287-146-8-1969 · Cited by 512
THE MAGIC BULLETS AND TUBERCULOSIS DRUG TARGETS
Zhang, Y. (2005) · Annual Review of Pharmacology and Toxicology · doi:10.1146/annurev.pharmtox.45.120403.100120 · Cited by 432
Metadata
- Authors
- Pelicic, V., Jackson, M., Reyrat, J.-M., Jacobs, W. R., Gicquel, B., & Guilhot, C.
- Venue
- Proceedings of the National Academy of Sciences
- Published
- 1997-09-30
- Volume
- 94
- Issue
- 20
- Pages
- 10955-10960
- ISSN
- 0027-8424 (ISSN-L)
- 1091-6490
- OpenAlex
- W2118508196
- Cited by
- 423
- References
- 0
- FWCI
- 17.76
- Type
- journal article
- Language
- en
- Open access
- green
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- Pelicic, V., Jackson, M., Reyrat, J.-M., Jacobs, W. R., Gicquel, B., & Guilhot, C. (1997). Efficient allelic exchange and transposon mutagenesis in Mycobacterium tuberculosis. Proceedings of the National Academy of Sciences. https://doi.org/10.1073/pnas.94.20.10955
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