Development of a Gene Replacement Method for the Filamentous Bacterium Leptothrix cholodnii SP-6
Tatsuki Kunoh ,
Erika Ono,
Tatsuya Yamamoto,
Ichiro Suzuki,
Minoru Takeda,
Nobuhiko Nomura
Affiliations
Tatsuki Kunoh
Faculty of Life and Environmental Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8577, Japan
Erika Ono
Graduate School of Life and Environmental Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8577, Japan
Tatsuya Yamamoto
Faculty of Life and Environmental Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8577, Japan
Ichiro Suzuki
Division of Materials Science and Chemical Engineering, Faculty of Engineering, Yokohama National University, 79-5 Tokiwadai, Hodogaya, Yokohama 240-8501, Japan
Minoru Takeda
Division of Materials Science and Chemical Engineering, Faculty of Engineering, Yokohama National University, 79-5 Tokiwadai, Hodogaya, Yokohama 240-8501, Japan
Nobuhiko Nomura
Faculty of Life and Environmental Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8577, JapanMicrobiology Research Center for Sustainability (MiCS), University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8577, Japan
Genetic strategies such as gene disruption and fluorescent protein tagging largely contribute to understanding the molecular mechanisms of biological functions in bacteria. However, the methods for gene replacement remain underdeveloped for the filamentous bacteria Leptothrix cholodnii SP-6. Their cell chains are encased in sheath composed of entangled nanofibrils, which may prevent the conjugation for gene transfer. Here, we describe a protocol optimized for gene disruption through gene transfer mediated by conjugation with Escherichia coli S17-1 with details on cell ratio, sheath removal, and loci validation. The obtained deletion mutants for specific genes can be used to clarify the biological functions of the proteins encoded by the target genes.Graphical overview
