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Targeted chromosomal knockouts in Mycoplasma pneumoniae.

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.

SUBMITTER: Krishnakumar R

PROVIDER: S-EPMC2916464 | biostudies-literature | 2010 Aug

REPOSITORIES: biostudies-literature

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Targeted chromosomal knockouts in Mycoplasma pneumoniae.

Krishnakumar Radha R Assad-Garcia Nacyra N Benders Gwynedd A GA Phan Quang Q Montague Michael G MG Glass John I JI

Applied and environmental microbiology 20100611 15

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 t ...[more]

PMID: 20543037

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