Illumina NovaSeq 6000 paired end sequencing; RNA-seq analysis of the fungal mycelium of the plant pathogen Pyricularia oryzae treated with and antimicrobial peptide analog of a Trichoderma peptaibol
RNA-seq analysis of the fungal mycelium of the plant pathogen Pyricularia oryzae treated with and antimicrobial peptide analog of a Trichoderma peptaibol
Conidia of P. oryzae (IT10 strain) were cultured in 1.8 mL Complete Medium (CM; Talbot et al., 1993) at a final concentration of 2x105 conidia per mL for 3 days at 30 °C under shaking at 150 rpm. After three days, 50 µM of Pep 4rink, one of the most effective peptides in vivo, were added to samples containing actively growing mycelium. Untreated samples were used as control. Three h after peptide treatment, fungal mycelia were collected, filtered with 100 µm Cell Strainer (Corning®), rinsed with sterile water and stored at -80 °C for RNA extraction. Total RNA was extracted with the RNeasy Plant Mini kit (Qiagen) following the manufacturer's instructions, performing the DNase digestion directly on the column. Two µg of total RNA were subjected to PolyA tail capture and rRNA depletion for selection of mRNA. Directional (stranded) RNA libraries were prepared, quantified and subjected to quality control. Sequencing was performed on Illumina Nova Seq 6000 platform.