Malt agar

The morphological characteristics of the fungus were studied by observing the color and texture of the colonies, as well as the color and shape of the sclerotia produced. Half cm diameter agar disk from 7-day-old fungal culture inoculated at the center of 9 cm diameter Petri dish using different culture media including Czapek-dox agar [19 ], Potato dextrose agar (PDA) [20 ], and Malt agar [21 ], then incubated for 7–10 days at 25 °C, all the used chemicals were purchased from (Sigma-Aldrich, St. Louis, MO, USA). Colony texture, the shape, and color of sclerotia were examined with the visual inspection and recorded after four to six days of incubation. Moreover, the fungus' microscopic characterization, including branching patterns and septation of mycelium, was also examined under a light microscope.

Aspergillus fumigatus strains were grown on Aspergillus minimal medium (AMM) agar plates (70 mM NaNO₃, 11.2 mM KH₂PO₄, 7 mM KCl, 2 mM MgSO₄, 1% [w/v] glucose and 1 µL/mL trace element solution at pH 6.5). The trace element solution was composed of 1 g of FeSO₄ • 7 H₂O, 8.8 g of ZnSO₄ • 7 H₂O, 0.4 g of CuSO₄ • 5 H₂O, 0.15 g of MnSO₄ • H₂O, 0.1 g of NaB₄O₇ • 10 H₂O, 0.05 g of (NH₄)₆Mo₇O₂₄ • 4 H₂O, and ultrafiltrated water to 1000 mL. Spores were harvested after 5 d (or 7 d for the evolution experiment) with 10 mL of sterile distilled water using a T-shaped inoculation spreader, and spore suspensions were filtered through a 30-µm cell strainer (MACS; Miltenyi Biotec GmbH) to exclude mycelium. A. fumigatus transformants were screened on AMM agar plates supplemented with 150 µg/mL hygromycin, 0.1 µg/mL pyrithiamine, or 160 µg/mL phleomycin for selection. For in vitro transcription and confocal laser scanning microscopy experiments, A. fumigatus AfS150 was grown on malt agar (Sigma-Aldrich) plates supplemented with agar to a final concentration of 2% (w/v) for 5 d. Escherichia coli strains were grown at 37°C on LB broth or agar supplemented with 100 µg/mL carbenicillin when necessary.