GSM8426827: mdx; Mus musculus; RNA-Seq
Lower hindlimbs of two mice were dissected, and dissociated with a collagenase B / dispase II solution with the gentleMACS Octo Dissociator with Heaters. Lysates were then filtered through a 100 μm filter and cells pelleted by centrifugation. Cell pellets were resuspended in red blood cell lysis buffer and washed with FACS buffer (5% FBS, 1 mM EDTA in PBS). After incubation with antibodies, MuSCs were sorted based on negative lineage markers (Cd11b−, Sca1−, Cd45−, Cd31−) and positive selection markers α7-integrin and VCAM1. Cell sorting was performed using a BD FACSAriaTM III (BD Biosciences). Cell were captured on the 10X Genomics Chromium platform and subjected to single cell transcriptomic sequencing. Following satellite cell isolation, the single-cell RNA-sequencing libraries were prepared using the 10X Genomics NextGen scRNA 3' V3.1 (10X Genomics, Pleasanton, CA) with approximately 10,000 satellite cells from each strain. Libraries were then sequenced with an Illumina NovaSeq6000 sequencer (Illumina, San Diego, CA).