title: A metabolic pathway for competitive saprophytic metabolism of chitin by terrestrial bacteria. RNA sequencing data
design: A metabolic pathway for competitive saprophytic metabolism of chitin by terrestrial bacteria. RNA sequencing data
library_construction_protocol: After sample growth (see growth protocol), 5 mL of each culture was pelleted, frozen in liquid nitrogen, and kept at -80°C before RNA extraction. S. venezuelae was grown in triplicate in 20 mL minimal glucose medium and 20 mL (10^6 spores / ml)  minimal chitin medium, both supplemented with (NH4)2(SO4) for 24 hours. 5 mL of each culture was pelleted, frozen in liquid nitrogen, and kept at -80°C before RNA extraction. S. venezuelae was grown in triplicate in 20 mL minimal glucose medium and 20 mL minimal chitin medium, both supplemented with (NH4)2(SO4) for 24 hours. 5 mL of each culture was pelleted, frozen in liquid nitrogen, and kept at -80°C before RNA extraction. RNA was extracted from the cell pellets using the Qiagen RNeasy Micro Kit (Qiagen, no. 74004) according to the manufacturer's protocol, with a minor modification: 0.5 mL of 0.5 mm glass beads were added to samples in lysis buffer followed by bead beating (six times one minute beating versus one minute on ice). RNA integrity was verified by gel electrophoresis before being sent for sequencing at Genome Quebec. Total RNA was quantified and its integrity was assessed using a LabChip GXII (PerkinElmer) instrument. rRNA was depleted from 250 ng of total RNA using QIAseq FastSelect (-5S/16S/23S Kit 96rxns). cDNA synthesis was achieved with the NEBNext RNA First Strand Synthesis and NEBNext Ultra Directional RNA Second Strand Synthesis Modules (New England BioLabs). Library preparation was then completed using the NEBNext Ultra II DNA Library Prep Kit for Illumina (New England BioLabs). Adapters and PCR primers were purchased from New England BioLabs. Libraries were quantified by Kapa Illumina GA with Revised Primers-SYBR Fast Universal kit (Kapa Biosystems). Average DNA fragment sizes were determined using a LabChip GXII (PerkinElmer) instrument. The libraries were normalized, pooled and then denatured in 0.05 N NaOH and neutralized using hybridization buffer (HT1).