Chromosomal Mutagenesis

Volume 435 of the series Methods in Molecular Biology pp 191-201

Allelic Exchange of Unmarked Mutations in Mycobacterium Tuberculosis

  • Martin S. PavelkaJr.Affiliated withDepartment of Microbiology and Immunology, University of Rochester Medical Center

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Mycobacterium tuberculosis has been studied since the 19th century, but genetic manipulation of this organism has only become possible within the last decade of the 20th century. One key methodology is the allelic exchange of unmarked, in-frame deletion mutations and point mutations using counter-selectable suicide plasmids. I describe below the challenges of allelic exchange in M. tuberculosis, the overall mechanism of a suicide vector system, and how it can be used for efficient allelic exchange in this formally intractable organism.

Key Words

Allelic exchange counter-selection electroporation Mycobacterium tuberculosis mutagenesis sacB sucrose