Suzuki, Hiroko Tokushima University
Shimokita, Eisuke Tokushima University|Tokushima Prefectural Agriculture, Forestry and Fisheries Technology Support Center Tokushima University Educator and Researcher Directory
Chiba, Hirofumi Tokushima University
Noji, Sumihare Tokushima University Tokushima University Educator and Researcher Directory KAKEN Search Researchers
Osakabe, Yuriko Tokushima University Tokushima University Educator and Researcher Directory KAKEN Search Researchers
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, CcDED1pro, 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 CcDED1pro 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.
Supplementary tables and figures : srep_7_1260_s1.pdf
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srep_7_1260.pdf 5.31 MB
srep_7_1260_s1.pdf 2.87 MB
Bioscience and Bioindustry