Scientific Reports (Apr 2017)

Genome editing in the mushroom-forming basidiomycete Coprinopsis cinerea, optimized by a high-throughput transformation system

  • Shigeo S. Sugano,
  • Hiroko Suzuki,
  • Eisuke Shimokita,
  • Hirofumi Chiba,
  • Sumihare Noji,
  • Yuriko Osakabe,
  • Keishi Osakabe

DOI
https://doi.org/10.1038/s41598-017-00883-5
Journal volume & issue
Vol. 7, no. 1
pp. 1 – 9

Abstract

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Abstract Mushroom-forming basidiomycetes produce a wide range of metabolites and have great value not only as food but also as an important global natural resource. Here, we demonstrate CRISPR/Cas9-based genome editing in the model species Coprinopsis cinerea. Using a high-throughput reporter assay with cryopreserved protoplasts, we identified a novel promoter, CcDED1 pro , with seven times stronger activity in this assay than the conventional promoter GPD2. To develop highly efficient genome editing using CRISPR/Cas9 in C. cinerea, we used the CcDED1 pro to express Cas9 and a U6-snRNA promoter from C. cinerea to express gRNA. Finally, CRISPR/Cas9-mediated GFP mutagenesis was performed in a stable GFP expression line. Individual genome-edited lines were isolated, and loss of GFP function was detected in hyphae and fruiting body primordia. This novel method of high-throughput CRISPR/Cas9-based genome editing using cryopreserved protoplasts should be a powerful tool in the study of edible mushrooms.