GSM2668035: Fermentor N°1 Sample 8h (D3); Clostridium beijerinckii; RNA-Seq
Cell lysis and RNA isolation was done with the TRIzol Plus RNA purification kit (AMbion). 5 mL TRIzol reagent is added to the frozen pellet followed by 1 mL chloroform after cell lysis. After shacking and centrifugation, the upper phase is mixed with an equal volume of 70% ethanol before transfer to PureLink RNA Spin Cartridges furnished with the kit. Kit instructions allow to obtain purified total RNA which is further processed by cleanup with the RNeasy mini kit (Qiagen).  DNA is eliminated after DNAse I treatment (AM1906, Invitrogen). 20µL RNA is treated with 2 µL DNAse in 50 µL nuclease free water for 30 min at 37°C before addition of another 1.5 µL DNAse and 30 min extra incubation. Treatment is done twice and is followed by RNA cleanup with the RNeasy kit. A full quality assessment of the RNA, quantification using Nanodrop, PCR to check for residual DNA and migration on 1% agarose gel is done before depletion of ribosomal RNA with MicrobExpress kit (ThermoFischer). Depletion uses 2-10 µg total RNA in 15 µL water. RNA libraries were prepared for sequencing using standard Illumina protocols
GEO Accession
GSM2668035