microRNA repertoire of wild-type C57BL/6 mice T cell blasts and their secreted small extracellular vesicles (Evs)

T cell blasts were obtained from wild-type C57BL/6 mice. Six-day culture blast supernatants were ultracentrifuged for small EV obtention and small EV pellet obtained after from 100,000g ultracentrifugation and T cell blasts RNA were isolated for small RNA sequencing; (N=3) independent experiments Total splenocytes were isolated from spleens of 8-to-12-week old WT C57BL/6 mice.Cells were cultured in the presence of Concanavalin and EVs were isolated from supernatants of 6-day lymphoblast cultures by serial ultracentrifugation. Total RNA from cells and secreted small EVs was isolated using the miRNeasy RNA Isolation Kit (QIAGEN). RNA was amplified and prepared for small RNA-Seq using the SmallRNA NEBNext Library Prep Set (New England Biolabs) following the manufacturer’s recommendations and sequenced using the Illumina HiSeq 2500 System. Three biological replicates were analyzed.