RNA-Seq of Fomes fomentarius
The RNA from the pure culture of F. fomentarius was extracted using a NucleoSpin RNA Plant kit (Macherey-Nagel) according to manufacturers protocol after mixing with 900 l of the RA1 buffer and shaking on FastPrep-24 (MP Biomedicals) at 6.5 ms1 twice for 20 s. Extracted RNA was treated as described previously (Tlskal et al. 2021) with OneStep PCR Inhibitor Removal kit (Zymo Research), DNA was removed using DNA-free DNA Removal Kit (Thermo Fisher Scientific). The efficiency of DNA removal was confirmed by the negative PCR results with the bacterial primers 515F and 806R (Caporaso et al. 2012). RNA quality was assessed using a 2100 Bioanalyzer (Agilent Technologies). rRNA in RNA samples was reduced using a Ribo-Zero rRNA Removal Kit Human/Mouse/Rat (Illumina). The efficiency of the removal was checked using a 2100 Bioanalyzer and removal was repeated when necessary. Reverse transcription was performed with SuperScript III (Thermo Fisher Scientific). Libraries for high-throughput sequencing were prepared using the ScriptSeq v2 RNA-Seq Library Preparation Kit (Illumina) according to the manufacturers instructions with a final 14 cycles of amplification by FailSafe PCR Enzyme (Lucigen).