GSM9531794: Scar, GR2-D0, scRNAseq; Homo sapiens; RNA-Seq
Skin biopsy samples (n=16) were thawed and washed with Dulbecco's Phosphate-Buffered Saline (DPBS) supplemented with 5% foetal calf serum (FCS) once before incubating with 1mg/ml collagenase P (Sigma Aldrich, UK) and 100mg/ml DNase I (Sigma Aldrich, UK) for 3.5 hours at 37°C in a humidified CO2 incubator. Cell suspensions were filtered through a 70mm cell strainer. Dissociated cells were then labelled with live and dead dye before live cell sorting using a BD LSR FORTESSAÔ X-20 Cell Analyzer (BD Biosciences, USA). Single cells were captured and barcoded using 10x Chromium platform and the sequencing libraries were prepared following manufacturer's instructions using dual index Chromium Next GEM G Single Cell 3' kit v3.1 (10xGemonics, California, USA) Single cells were captured and barcoded using 10x Chromium platform and the sequencing libraries were prepared following manufacturer's instructions using dual index Chromium Next GEM G Single Cell 3' kit v3.1 (10xGemonics, California, USA). Libraries were quantified using an Agilent Bioanalyzer High Sensitivity DNA chip and pooled together to get similar numbers of reads from each single cell before sequencing. Overall, approximately 50,000 cells per sample were used for construction libraries and sequenced using Novogene single cell RNA sequencing services.