Novel insights into the role of the histone variant H2A.Z in the plant pathogen Fusarium fujikuroi [RNA-seq]

Transcriptome in respect to H2A.Z downregulation or overexpression The F. fujikuroi IMI58289 wild-type strain (FfWT) provided by Commonwealth Mycological Institute, Kew, UK, was used as a parental strain to generate the H2A.Z knock-down strain (Tetoff::FfH2A.Z), the constitutive H2A.Z overexpression (OE::FfH2A.Z) strain as well as the N- and C-terminal hemagglutinin (HA)-tagged H2A.Z (HA::FfH2A.Z, FfH2A.Z::HA) strains. Submerged fungal growth was performed in 100 mL Darken pre-culture medium (Darken et al., 1959) on a rotary shaker at 180 rpm, 30°C in constant darkness for 3 days. Cultures inoculated with the TetOff::H2A.Z knock down strain were additionally supplemented with 0-50 µg/mL DOX to facilitate FfH2A.Z gene silencing. Flasks were incubated on a rotary shaker (180 rpm, 30°C) in the dark. Mycelia were harvested 3-4 days post inoculation (dpi) for RNA isolation. Isolation of fungal RNA was performed with lyophilized and ground mycelia from cultivation on solid CM or under liquid-standard conditions (ICI supplemented with 6 or 60 mM glutamine) for 3-4 days. Total RNA was extracted from samples, using the RNA reagent TRIzol (Thermo Fisher Scientific) as described in the manufacturer’s manual.