Evaluation of CRISPR/Cas9 system for transformation of N. crassa. a Diagram of donor plasmids. Blue color regions are the sequence regions that are homologous to genomic DNA. Bar gene cassette which contains the bar gene under the control of trpC promoter and terminator was included in each plasmids for selection. b The number of Ignite-resistant colonies by clr-2 locus targeted transformation with different amount of Cas9 and gRNA plasmids. From left to right: zero, 1, 2.5, and 5 µg each of Cas9 and gRNA plasmid was co-transfected with the donor plasmid (5 µg). **p < 0.01, Tukey’s test. Error bars corresponds to the SEM. c The number of Ignite-resistant colonies by csr-1 locus targeted transformation with zero and 5 µg each of Cas9 and gRNA plasmids. **p < 0.01, student’s t-test. Error bars corresponds to the SEM. d–f Luciferase activity on the plates of gsy-1-luciferase transformants. Luciferase signal (d), under red-light (e), and merged (f) images are shown.