title: Development of a novel plasmid vector pTIO-1 adapted for electrotransformation of Porphyromonas gingivalis.
abstract: We report here the construction of a plasmid vector designed for the efficient electrotransformation of the periodontal pathogen Porphyromonas gingivalis. The novel Escherichia coli-Bacteroides/P. gingivalis shuttle vector, designated pTIO-1, is based on the 11.0-kb E. coli-Bacteroides conjugative shuttle vector, pVAL-1 (a pB8-51 derivative). To construct pTIO-1, the pB8-51 origin of replication and erythromycin resistance determinant of pVAL-1 were cloned into the E. coli cloning vector pBluescript II SK(-) and non-functional regions were deleted. pTIO-1 has an almost complete multiple cloning site from pBluescript II SK(-). The size of pTIO-1 is 4.5kb, which is convenient for routine gene manipulation. pTIO-1 was introduced into P. gingivalis via electroporation, and erythromycin-resistant transformants carrying pTIO-1 were obtained. We characterized the transformation efficiency, copy number, host range, stability, and insert size capacity of pTIO-1. An efficient plasmid electrotransformation of P. gingivalis will facilitate functional analysis and expression of P. gingivalis genes, including the virulence factors of this bacterium.
authors.affiliation: Department of Oral Microbiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University, Okayama 700-8558, Japan; Department of Orthodontics, Okayama University Hospital, Okayama 700-8558, Japan.
authors.forename: Junpei
authors.initials: J
authors.lastname: Tagawa
authors.affiliation: Department of Oral Microbiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University, Okayama 700-8558, Japan; Advanced Research Center for Oral and Craniofacial Sciences, Dental School, Okayama University, Okayama 700-8558, Japan.
authors.forename: Tetsuyoshi
authors.initials: T
authors.lastname: Inoue
authors.affiliation: Division of Microbiology and Oral Infection, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Nagasaki University, Nagasaki 852-8588, Japan.
authors.forename: Mariko
authors.initials: M
authors.lastname: Naito
authors.affiliation: Division of Microbiology and Oral Infection, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Nagasaki University, Nagasaki 852-8588, Japan.
authors.forename: Keiko
authors.initials: K
authors.lastname: Sato
authors.affiliation: Department of Molecular Microbiology, Faculty of Medicine, Kagawa University, Kagawa 761-0793, Japan.
authors.forename: Tomomi
authors.initials: T
authors.lastname: Kuwahara
authors.affiliation: Department of Oral Microbiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University, Okayama 700-8558, Japan; Advanced Research Center for Oral and Craniofacial Sciences, Dental School, Okayama University, Okayama 700-8558, Japan.
authors.forename: Masaaki
authors.initials: M
authors.lastname: Nakayama
authors.affiliation: Division of Microbiology and Oral Infection, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Nagasaki University, Nagasaki 852-8588, Japan.
authors.forename: Koji
authors.initials: K
authors.lastname: Nakayama
authors.affiliation: Department of Orthodontics and Dentofacial Orthopedics, Graduate School of Dentistry, Osaka University, Osaka 565-0871, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Yamashiro
authors.affiliation: Department of Oral Microbiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University, Okayama 700-8558, Japan; Advanced Research Center for Oral and Craniofacial Sciences, Dental School, Okayama University, Okayama 700-8558, Japan. Electronic address: oharan@md.okayama-u.ac.jp.
authors.forename: Naoya
authors.initials: N
authors.lastname: Ohara
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001439:Bacteroides / Q000235:genetics; D024881:Drug Resistance, Bacterial; D018274:Electroporation; D004917:Erythromycin / Q000494:pharmacology; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D005827:Genetics, Microbial / Q000379:methods*; D042822:Genomic Instability; D058507:Host Specificity; D006801:Humans; D008967:Molecular Biology / Q000379:methods*; D010957:Plasmids; D016966:Porphyromonas gingivalis / Q000235:genetics*; D018741:Replication Origin; D012641:Selection, Genetic; D014169:Transformation, Bacterial
chemical_list: D000900:Anti-Bacterial Agents; D004917:Erythromycin
keywords: Electroporation; Periodontal disease; Plasmid; Porphyromonas; Transformation