GSM6199122: BMDMB2-WT-Rep3; Mus musculus; RNA-Seq
Total RNA was isolated using the PureLink™ RNA Mini Kit (Invitrogen, 12183025). RNA sequencing was performed by the University of Texas Genomic Sequencing and Analysis Facility. Poly(A) enrichement was performed.  mRNA was isolated from total RNA using the Poly(A) Purist-MAG kit (ThermoFisher). Briefly, total RNA was combined in a 96-well plate with washed and resuspended Oligo (dT) MagBeads. The mixture was heated in a thermocycler set to 70°C for 5 minutes, followed by incubation at room temperature with gentle vortexing for 30 minutes. Subsequently, the beads were captured and washed, and bound poly(A) RNA was eluted in water.  mRNA quality was assessed on the Agilent Bioanalyzer using the Agilent RNA 6000 Pico kit (Agilent). Libraries were prepared at the University of Texas Genomic Sequencing and Analysis Facility according to manufacturer's instructions for the NEBNext Ultra II Directional RNA kit (NEB, product number E7760). The resulting libraries tagged with unique dual indices were checked for size and quality using the Agilent High Sensitivity DNA Kit (Agilent). Library concentrations were measured using the KAPA SYBR Fast qPCR kit and loaded for sequencing on the NovaSeq 6000 instrument (paired end 2X150).