Fig. 5

Verticillium dahliae VdZFP1 and VdZFP2 regulate the melanization of microsclerotia. A Microsclerotia morphology of the WT, ΔVdZFP1, EC^ΔVdZFP1, ΔVdZFP2, EC^ΔVdZFP1, and ΔVdZFP1_2 strains exposed to the melanin biosynthesis pathway inhibitor tricyclazole and intermediate scytalone. All of the indicated strains were cultured on the BMM medium or supplemented with the tricyclazole (60 mg/mL) and scytalone (50 mg/mL) for 7 and 14 days. The microsclerotia were observed using a stereoscope, scale bar = 100 μm. B Microsclerotia phenotypes of albino ΔVdCmr1 and double-deleted mutants (ΔVdZFP1_VdCmr1 and ΔVdZFP2_VdCmr1). The microsclerotia were inducted on BMM medium for 7 and 14 days and were observed using a stereoscope, scale bar = 100 μm. C Investigation of microsclerotia melanization in ΔVdPKS1 and double-deleted mutants (ΔVdZFP1_VdPKS1 and ΔVdZFP2_VdPKS1) supplemented with the melanin intermediate scytalone. The microsclerotia were inducted on normal or scytalone (50 mg/mL) supplemented BMM medium for 7 days and were observed using a stereoscope, scale bar = 100 μm. D Relative expression analyses of the melanin biosynthesis genes of the indicated strains in C following supplementation with scytalone. The strains cultured on BMM medium contained 50 mg/mL scytalone for 7 days were collected from above (C) experiment. Using ΔVdPKS1 as control, the expression levels of each gene in mutants were detected by RT-qPCR for 3 repetitions using the 2^−ΔΔCT method. Error bars represent standard errors of the mean, and ^*P < 0.05, ^**P < 0.01, and ^***P < 0.001 (one-way ANOVA). All experiments were independently repeated 3 times