title: NextSeq 550 sequencing; Effect of Hst3p inhibition in Candida albicans: a genome-wide H3K56 acetylation analysis
design: Effect of Hst3p inhibition in Candida albicans: a genome-wide H3K56 acetylation analysis
library_construction_protocol: As described above, C. albicans cultures were grown with or without 10 mM NAM in Petri dishes of 90 mm diameter and incubated at 25°C. Three independent biological replicates were performed for either CTRL or CaNAM cells. After 28 h of incubation, a total of 109 yeast cells were harvested (8,000g, 10 min at 4°C) and washed with diethylpyrocarbonate (DEPC)-treated water. RNA isolation was performed as described by Vennapusa et al., 2020 with some modifications. Briefly, yeast cells were resuspended in 600 mL of RNA extraction buffer (100 mM Tris- HCl (pH: 8), 25 mM EDTA, PVP 2.5%, NaCl 2.5 M, 2.5 % β-Mercaptoethanol, 2% SDS) and disrupted mechanically with BeadBug microtube homogenizer (Benchmark Scientific, Sigma-Aldrich, Milan, Italy) by using acid-washed glass beads. Total RNA was isolated using an equal volume of Acid-Phenol/Chloroform, pH 4.5 with IAA, 125:24:1 (Sigma-Aldrich, Milan, Italy). Following centrifugation, the RNA was precipitated from the aqueous phase with 100% ethanol, washed twice with 75% ethanol, and dissolved in DEPC-water. RNA quantification was carried out with the instrument Nanodrop 200 Thermo Fisher Scientific (Monza, Italy), and its integrity was checked by electrophoresis on 1.5% agarose gel. For RNA sequencing, RNA quality was assessed with TapeStation (Agilent, Cernusco sul Naviglio, Italy), and only RNA with RIN > 8 was used for library production.