GSM4057044: RNA_seq_SingleCell_dpp40; Mus musculus; RNA-Seq
Subcutaneous fat was removed from the skin with a scalpel, and the whole skin was trypsinized at 37 °C for an hour to remove the dermis. The cell suspensions were filtered through strainers (70-μm, BD Falcon), collected by centrifugation (300 × g, 5 min) and suspended in Dulbecco's modified Eagle's medium (Ca−). Cell suspensions were incubated with Pacific Blue anti-human/mouse CD49f (Biolegend) antibody and eFluor660 anti-mouse CD34 (Biolegend) antibody for 30 min on ice, washed twice with PBS and subjected to flow cytometric analysis. Cell isolation was performed on a FACSAria Ⅲ sorter. RNA was extracted from CD49f positive and CD34 negative cells using the RNeasy Micro Kit (Qiagen). Quality RNA samples were selected (RIN>7).  For scRNAseq, single cell suspensions of all samples were resuspended in PBS-1% BSA at 1000cells/μL and barcoded with a 10x Chromium Controller (10x Genomics). RNA-seq libraries were generated using the TruSeq Stranded mRNA library prep kit (Illumina) and the libraries were sequenced on NextSeq 500 (Illumina). For scRNAseq, RNA from the barcoded cells for each sample was subsequently reverse-transcribed and sequencing libraries were constructed with reagents from a Chromium Single Cell v2 reagent kit (10x Genomics). Sequencing was performed with Illumina NextSeq 500 .
GEO Accession
GSM4057044