GSM8329705: sgASH1L_2; Mus musculus; RNA-Seq
Control and ASH1L-knockout metastatic bone tumors were dissected, minced, and digested into single-cell suspension using Collagenase A (Sigma, 11088793001) at 37°C for 45min with continuously gentle shaking. The dissociated tissues were centrifuged at 500xg for 5 min, followed by further digestion with 0.25% Trypsin (Corning, 25053CI) at 37°C for 5 min. The samples were filtered using a 70uM strainer (Falcon, 352350) and centrifuged at 500xg at 4°C for 5 min, followed by incubation in 1x RBC lysis buffer (Biolegend, 420302) at RT for 5 min. After washing with cold DMEM, the cells were collected and resuspended in cold PBS containing 0.04% BSA (Ambion, AM2616), followed by measurement of cell number and viability. The scRNA-seq libraries were generated using the 10X Genomics Chromium Controller Instrument and Chromium Single Cell 30 V3 Reagent Kits (10X Genomics) according to the manufacturer's recommendations. Libraries were subsequently sequenced on NovaSeq 6000 (Illumina) with a paired-end 50-base pair (50PE) reading strategy.