Bars are the mean and SD of five replications. Differences between wild type and the mutants

were found significant according to t-test (P < 0.05) in the following treatments: sporulation in the light (A), sporulation in dark on EMS with 500 μM IAA (B, C), sporulation in the dark on EMS with 250 μM (C). Repetition of experiments led to similar results. Next we tested the possible effect of IAA on sporulation. Wild-type and mutant strains were cultured on media with 500 μM IAA. The plates were kept in the dark to prevent photo-oxidation of IAA, and to eliminate light-induced differences in sporulation between the wild type and mutants. IAA significantly enhanced sporulation in wild-type cultures under these conditions, Fosbretabulin datasheet while it had no effect on sporulation of the cgopt1-silenced mutants (Fig. 6B). Furthermore, the effect of IAA on sporulation in wild-type cultures was dose-dependent: a small increase in spore production see more was observed at 100 μM IAA, and production was further enhanced by 250 μM and 500 μM IAA (Fig. 6C). No change was observed in the sporulation of the mutants, regardless

of IAA concentration. These results showed a clear and consistent phenotype caused by IAA, which is abolished in the cgopt1-silenced mutants. Colony morphology While characterizing the transcriptional response to IAA, we noticed the development of more compact mycelium in the presence of auxin. To further examine this phenotype, we tested the effect of IAA on the development of mycelia in liquid culture. In REG medium, the wild-type colonies

accumulated intense orange pigmentation, while the silenced mutants developed a very pale orange color Megestrol Acetate (Fig. 7, top). This phenotype was similar to that observed on solid REG plates (Fig. 5A). IAA greatly reduced pigmentation in wild-type cultures, whereas it had no effect on the mutants, which retained their light orange color (Fig. 7, top). Figure 7 Effect of culture media and IAA on morphology of wild type and cgopt1 mutants. Similar results were obtained with Ori51 and Ori83 mutant strains. Only the results with Ori51 are presented. Top: Colonies of wild type and Ori51 mutant strain grown in REG liquid medium for 3 days in the absence (-) and presence (+) of 500 μM IAA. Middle: stereoscope images of individual pellets that developed in REG media with or without 500 μM IAA. Bottom: stereoscope images of individual pellets that developed in CD medium with or without 500 μM IAA. Bar = 1 mm. A difference was noted in the morphology of wild-type and mutant colonies. In REG medium, the wild type developed pellets surrounded by long hyphae, which became more compact with shorter hyphae when IAA was added (Fig. 7, middle). Under these conditions, the cgopt1-silenced mutants developed compact pellets without free hyphae, and this morphology did not change in the presence of IAA.