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Construction of a plasmid interspecific transfer system in Bacillus species with the counter-selectable marker mazF

  • Biotechnology Methods - Original Paper
  • Published:
Journal of Industrial Microbiology & Biotechnology

Abstract

Bacillus sp. strains as attractive hosts for the production of heterologous secretory proteins usually play important roles in bio-industry. However, low transformation efficiency of exogenous plasmids limited the application of Bacillus species. Here, a novel plasmid interspecific transfer system, with high transformation efficiency, high positive rate, and convenient manipulation, has been successfully constructed. A high electrotransformation efficiency strain Bacillus subtilis F-168 containing the counter-selectable marker mazF was used as the plasmid donor strain in this transfer method. A shuttled plasmid pBE980 and its recombinant plasmids pBE980::pulA and pBE980::HSPA were successfully transferred into the recipient Bacillus strains (Bacillus amyloliquefaciens 66, Bacillus licheniformis 124 and Bacillus megaterium 258) by this method. After co-culturing the donor cells (OD600nm = 1.3–1.7) and the recipient cells (OD600nm = 0.5–0.9) for 24 h in 22 °C, more than 1.0 × 105 positive transformants were obtained and a interspecific transformation efficiency of 1.0 × 10−3. It would provide a new approach for genetic manipulation in Bacillus strains and accelerate the research progress of the wild Bacillus strains in bio-industry.

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Acknowledgements

This work was supported by the National Natural Science Fund of China (Grant 31701534), the Key Deployment Project in Chinese Academy of Sciences (Grant KFJ-STS-ZDTP-016-1, KFZD-SW-211-2), and the Tianjin Science & Technology Planning Project (Grant 16YFZCSY00790,16YFXTSY00530 and 15YFYssy00040).

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Author notes

  1. Xingya Zhao and Jianyong Xu contributed equally to this work.

Authors and Affiliations

  1. University of Chinese Academy of Sciences, Beijing, 100049, China

    XingYa Zhao, ZhenXiao Yu, ShiBin Yang & Hui Song

  2. Industrial Enzymes National Engineering Laboratory, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, China

    XingYa Zhao, JianYong Xu, Ming Tan, ZhenXiao Yu, ShiBin Yang, HongChen Zheng & Hui Song

  3. Tianjin Key Laboratory for Industrial Biological, Systems and Bioprocessing Engineering, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, China

    JianYong Xu, Ming Tan, HongChen Zheng & Hui Song

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  1. XingYa Zhao
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  2. JianYong Xu
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  4. ZhenXiao Yu
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  7. Hui Song
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Correspondence to HongChen Zheng or Hui Song.

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This article does not contain any studies with human participants or animals performed by any of the authors.

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The authors declare that they have no competing interests.

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Zhao, X., Xu, J., Tan, M. et al. Construction of a plasmid interspecific transfer system in Bacillus species with the counter-selectable marker mazF. J Ind Microbiol Biotechnol 45, 417–428 (2018). https://doi.org/10.1007/s10295-018-2038-0

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  • DOI: https://doi.org/10.1007/s10295-018-2038-0

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