Detailed Protocols for Studying Magnaporthe oryzae Growth and Virulence

M. oryzae strain Guy11 was used as wild type throughout this work. Both Guy11 and its derivative mutants were cultured on complete medium (CM) [89] (link) for 3–15 days at 28°C to assess the growth and colony characteristics. Fungal mycelia were harvested from liquid CM and used for genomic DNA and RNA extractions. To observe the vegetative growth under the oxidative stress condition, H₂O₂ (Aldrich, 323381, 3 wt. %) was mixed in solid CM, and diameters of fungal colonies were measured after 3 to 5 days as indicated. For the activation of the laccase activity in the Moap1 mutants, copper sulphate was amended in the CM medium for 1 mM at final concentration. Cell wall integrity assay was performed by growing strains in Congo Red (CR, Aldrich, 860956) amended CM medium (200 µg/ml) for 5 days. For conidia collection, strains were normally maintained on corn meal (RDC) medium [43] (link) at 28°C for 10 days and then transferred to constant fluorescent light condition to promote conidiation for another 3–5 days. Conidia were obtained by rubbing mycelia with water followed by filtration through Miracloth (Calbiochem, San Diego, USA). For mycelial growth assay, strains were inoculated in the liquid complete medium for 48 hrs and then transferred to the Petri dish for photograph. To observe conidiophore development and conidiation, strains were inoculated on RDC for 5 days and mycelia were rubbed with a glass rod before transferring to the constant fluorescent light condition to promote conidiation for another 2 days. S. cerevisiae strains were grown in SD medium supplemented with appropriate amino acids and with glucose (3% (w/v)) or galactose (2% (w/v)). All growth assays were repeated for three times, with three replicates each time.