Illumina NovaSeq 6000 paired end sequencing; RNA-seq of in vitro derived B cells
RNA-seq of in vitro derived B cells
Mouse splenic B cells were purified by negative selection and cultured in complete medium in the presence of in vitro differentiation cocktails (anti-IgM, R848, anti-CD40, IL-21 plus IFN-γ or IL-4) for 3 days. The in vitro dereived B cells were collected for RNA sequencing. The in vitro indued B cells were resuspended in TRIzol reagent for total RNA extraction. RNA integrity was assessed using the Fragment Analyzer 5400 (Agilent Technologies, CA, USA). Then total RNA was used as input material for the RNA sample preparations. Sequencing libraries were generated using NEBNext® UltraTM RNA Library Prep Kit for Illumina® (NEB, USA) following manufacturer's recommendations. The library fragments were purified with AMPure XP system (Beckman Coulter, Beverly, USA) and library quality was assessed on the Agilent Bioanalyzer 2100 system.