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Transformation by complementation of a uracil auxotroph of the hyper lignin-degrading basidiomycete Phanerochaete sordida YK-624

Applied Microbiology and Biotechnology volume 76, pages 1079–1091 (2007)Cite this article

Abstract

Phanerochaete sordida YK-624 is a hyper lignin-degrading basidiomycete possessing greater ligninolytic selectivity than either P. chrysosporium or Trametes versicolor. To construct a gene transformation system for P. sordida YK-624, uracil auxotrophic mutants were generated using a combination of ultraviolet (UV) radiation and 5-fluoroorotate resistance as a selection scheme. An uracil auxotrophic strain (UV-64) was transformed into a uracil prototroph using the marker plasmid pPsURA5 containing the orotate phosphoribosyltransferase gene from P. sordida YK-624. This system generated approximately 50 stable transformants using 2 × 107 protoplasts. Southern blot analysis demonstrated that the transformed pPsURA5 was ectopically integrated into the chromosomal DNA of all transformants. The enhanced green fluorescent protein (EGFP) gene was also introduced into UV-64. The transformed EGFP was expressed in the co-transformants driven by P. sordida glyceraldehyde-3-phosphate dehydrogenase gene promoter and terminator regions.

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Acknowledgments

This work was supported by grants-in-aid (Biodesign Program) from the Ministry of Agriculture, Forestry, and Fisheries of Japan, and Scientific Research (B) from the Ministry of Education, Science, Sports, and Culture of Japan.

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Affiliations

  1. National Agricultural Research Center for the Tohoku Region, National Agriculture and Food Research Organization, Arai, Fukushima, 960-2156, Japan

    Kenji Yamagishi, Toshiyuki Kimura & Sigeru Oita

  2. Faculty of Agriculture, Shizuoka University, Ohya 836, Suruga-ku, Shizuoka, 422-8529, Japan

    Tatsuki Sugiura & Hirofumi Hirai

Authors
  1. Kenji Yamagishi
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  2. Toshiyuki Kimura
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  3. Sigeru Oita
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  4. Tatsuki Sugiura
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  5. Hirofumi Hirai
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Correspondence to Kenji Yamagishi.

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Yamagishi, K., Kimura, T., Oita, S. et al. Transformation by complementation of a uracil auxotroph of the hyper lignin-degrading basidiomycete Phanerochaete sordida YK-624. Appl Microbiol Biotechnol 76, 1079–1091 (2007). https://doi.org/10.1007/s00253-007-1093-7

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  • DOI: https://doi.org/10.1007/s00253-007-1093-7

Keywords

  • Phanerochaete sordida
  • EGFP
  • Genetic transformation
  • PsURA5
  • Uracil auxotroph