A novel method for transforming the thermophilic bacterium Geobacillus kaustophilus

• Miyano, Megumi

• 田中, 耕生

• 石川, 周

• Mori, Kotaro

• Miguel-Arribas, Andres

• Meijer, Wilfried J. J.

• 吉田, 健一

抄録

Background: Bacterial strains of the genus Geobacillus grow at high temperatures of 50-75 °C and could thus be useful for biotechnological applications. However, genetic manipulation of these species is difficult because the current techniques for transforming Geobacillus species are not efficient. In this study, we developed an easy and efficient method for transforming Geobacillus kaustophilus using the conjugative plasmid pLS20cat. Results: We constructed a transformation system comprising (i) a mobilizable Bacillus subtilis-G. kaustophilus shuttle plasmid named pGK1 that carries the elements for selection and replication in Geobacillus, and (ii) a pLS20cat-har-boring B. subtilis donor strain expressing the dam methylase gene of Escherichia coli and the conjugation-stimulating rap(LS20) gene of pLS20cat. This system can be used to efficiently introduce pGK1 into G. kaustophilus by mobilization in a pLS20cat-dependent way. Whereas the thermostable kanamycin marker and Geobacillus replication origin of pGK1 as well as expression of dam methylase in the donor were indispensable for mobilization, ectopic expression of rap(LS20) increased its efficiency. In addition, the conditions of the recipient influenced mobilization efficiency: the highest mobilization efficiencies were obtained using recipient cells that were in the exponential growth phase. Furthermore, elimination of the origin of transfer from pLS20cat enhanced the mobilization. Conclusions: We describe a novel method of plasmid mobilization into G. kaustophilus recipient from B. subtilis donor depending on the helper function of pLS20cat, which enables simple, rapid, and easy transformation of the thermophilic Gram-positive bacterium.

収録刊行物

• Microbial Cell Factories

  Microbial Cell Factories 17 127-127, 2018-08-17

  BMC

キーワード

• Geobacillus kaustophilus
• Bacillus subtilis
• Plasmid
• Conjugation
• Mobilization
• Transformation

詳細情報

• CRID: 1050012570393331200
• NII論文ID: 120006510197
• ISSN: 14752859
• HANDLE: 20.500.14094/90005177
• 本文言語コード: en
• 資料種別: journal article
• データソース種別:
  • IRDB
  • CiNii Articles