GSM2360594: 2011_0026; Trichoderma reesei; RNA-Seq
At each time point, 5 mL culture medium containing mycelia was filtrated on 1.2 µL GF/C glass microfiber membranes and flash frozen in liquid nitrogen. Frozen mycelia were disrupted and homogenized in Lysing Matrix C (Fast RNA Pro Red Kit, MP Bio) in the presence of 700 µL Lysis buffer RLT (RNeasy Mini Kit, Qiagen) complemented with 7 µL of β-mercaptoethanol on a rotor-stator homogenizer (FastPrep, MP Bio, Santa Ana, CA, USA). After centrifugation, the lysate was transferred on a Qiashredder spin column (Qiagen) and centrifuged for 2 min. to eliminate all cell debris. The flow-through was then mixed with 0.5 volumes of pure ethanol and transferred to an RNeasy spin column. All further steps, including an on-column DNase digestion, were realized following the manufacturer’s instructions. RNA quantification was performed with Qubit 2.0 (Life Technologies, Carlsbad, CA, USA). The quantity and quality of the total RNA was determined using a NanoDrop ND-1000 spectrophotometer (Nanodrop Technologies, Wilmington, USA) and by electrophoresis on 1 % agarose gel. An additional quality control was performed on the Bioanalyzer 2100 system (Agilent, Santa Clara, USA). Library preparation and Illumina sequencing were performed at the Ecole normale superieure Genomic Platform (Paris, France). Messenger (polyA+) RNAs were purified from 1 µg of total RNA using oligo(dT) primer. Libraries were prepared using the strand non-specific RNA-Seq library preparation TruSeq RNA Sample Prep Kits v1 (Illumina). Libraries were multiplexed by 4 on one single flowcell lane and subjected to 50 bp paired-end read sequencing on a HiSeq 2000 device
GEO Accession
GSM2360594