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Appl Environ Microbiol. 2010 Aug;76(15):5297-9. doi: 10.1128/AEM.00024-10. Epub 2010 Jun 11.

Targeted chromosomal knockouts in Mycoplasma pneumoniae.

Author information

1
J. Craig Venter Institute, 9704 Medical Center Drive, Rockville, MD 20850, USA. rkrishna@jcvi.org

Abstract

Most gene knockouts in mycoplasmas are achieved through labor-intensive transposon mutagenesis. Here, we describe a method for making targeted deletions in Mycoplasma pneumoniae by use of homologous recombination. In this method, M. pneumoniae is transformed with a plasmid carrying an antibiotic resistance marker flanked by 1-kb regions surrounding the target gene. Following selection for the antibiotic resistance, colonies are screened for double crossovers which indicate complete deletion of the target open reading frame.

PMID:
20543037
PMCID:
PMC2916464
DOI:
10.1128/AEM.00024-10
[Indexed for MEDLINE]
Free PMC Article

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