GSM5570285: e135 caudal MNs; Mus musculus; RNA-Seq
Spinal cordssegments were dissected in L15 medium and cells were dissociated using enzymatic (papain) and mechanical approach, following manufacturer's instruction (Neural Tissue Dissociation Kit (P) and gentleMACS dissociator, Miltenyi Biotec). Cell suspension was filtered through 70µm strainer and subjected to FACS with 85um nozzle diameter under 45 psi to enrich GFP-positive cells. Collected cells were centrifuged at 1000rpm for 5 minutes and resuspended in 1%FBS supplemented DMEM. Cells viability and concentration was determined using Countess II Automated Cell Counter. Cells were loaded into 10x Chromium Controller using Single Cell 3' v2 reagents for single cell isolation. scRNA-seq libraries were generated following manufacturer's instruction (10X Genomics, Chromium Single Cell 3' v2) and sequenced using a NextSeq 500 platform with pair-ended reads (PE150: Read 1: 28 bp; Read 2: 122 bp) and NovaSeq S4 150 pair-ended reads (PE150: Read 1: 75 bp; Read 2: 75 bp).