GSM5353708: mESC +5uM IdU Bulk Rep 3; Mus musculus; RNA-Seq
For scRNA-seq, cells were trypsinized with TrypLE and spun down for 5 minutes at 200 x g. Single-cell suspensions were prepared in DPBS at a concentration of 83,000 cells/ml. Approximately 3000 cells from each sample were loaded into a chip and processed with the Chromium Single Cell Controller (10x Genomics). For mESC bulk RNA-seq, cells were trypsinized with TrypLE and RNA was extracted using a RNeasy minikit (Qiagen) according to manufacturer's instructions. ERCC spike-in RNA (2ml diluted at 1:100) was added to each RNA extraction (Ambion, cat:4456740). For MEF bulk RNA-seq, RNA was extracted from frozen cell pellet using a RNeasy minikit (Qiagen) according to manufacturer's instructions. To generate single-cell gel beads in emulsion (GEMs), samples were assigned unique indexes using Single Cell 3' Library and Gel Bead Kit V2 (10x Genomics, cat:120237). Sequencing was performed on an Illumina for 10x libraries. For bulk RNA-seq, cDNA libraries were prepared with an NEBNext Ultra II RNA Library Prep kit (NEB, cat:E7770S) and sequenced with an Illumina HiSeq4000.