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Development of a novel plasmid vector pTIO-1 adapted for electrotransformation of Porphyromonas gingivalis.
Tagawa, Junpei; Inoue, Tetsuyoshi; Naito, Mariko; Sato, Keiko; Kuwahara, Tomomi; Nakayama, Masaaki; Nakayama, Koji; Yamashiro, Takashi; Ohara, Naoya.
Afiliação
  • Tagawa J; 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.
  • Inoue T; 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.
  • Naito M; Division of Microbiology and Oral Infection, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Nagasaki University, Nagasaki 852-8588, Japan.
  • Sato K; Division of Microbiology and Oral Infection, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Nagasaki University, Nagasaki 852-8588, Japan.
  • Kuwahara T; Department of Molecular Microbiology, Faculty of Medicine, Kagawa University, Kagawa 761-0793, Japan.
  • Nakayama M; 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.
  • Nakayama K; Division of Microbiology and Oral Infection, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Nagasaki University, Nagasaki 852-8588, Japan.
  • Yamashiro T; Department of Orthodontics and Dentofacial Orthopedics, Graduate School of Dentistry, Osaka University, Osaka 565-0871, Japan.
  • Ohara N; 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.oka
J Microbiol Methods ; 105: 174-9, 2014 Oct.
Article em En | MEDLINE | ID: mdl-25102110
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.
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Texto completo: 1 Base de dados: MEDLINE Assunto principal: Transformação Bacteriana / Técnicas de Transferência de Genes / Eletroporação / Porphyromonas gingivalis / Vetores Genéticos / Genética Microbiana / Biologia Molecular Limite: Humans Idioma: En Revista: J Microbiol Methods Ano de publicação: 2014 Tipo de documento: Article País de afiliação: Japão

Texto completo: 1 Base de dados: MEDLINE Assunto principal: Transformação Bacteriana / Técnicas de Transferência de Genes / Eletroporação / Porphyromonas gingivalis / Vetores Genéticos / Genética Microbiana / Biologia Molecular Limite: Humans Idioma: En Revista: J Microbiol Methods Ano de publicação: 2014 Tipo de documento: Article País de afiliação: Japão