GSM9669436: GrnKO; Rosa26; 11; Mus musculus; RNA-Seq
RNA was harvested in RLT buffer with B-mercaptoethanol, collected into low-bind eppendorfs, and stored at -80oC until use. For each of the 16 biological samples, a sequencing library was prepared with the QuantSeq 3′ mRNA-Seq V2 Library Prep Kit FWD (Lexogen 193.384). The UMI second strand synthesis module was used to introduce unique molecular identifiers (UMIs), following the protocol defined by the manufacturer.  Briefly, oligo dT primers were hybridized to total RNA for reverse transcription, followed by RNA removal after first strand synthesis. UMIs were introduced during second strand synthesis and cDNA was purified using magnetic beads, followed by 18 cycles of PCR with dual indexes and PCR purification. Library concentration and fragment size distribution were assessed using the Agilent TapeStation D1000 Screentape system (5067-5582). Libraries were pooled in equimolar ratios and sequenced across two lanes of an Illumina NovaSeq X Series 10B flowcell.