GSM7157924: Healthy control skin patient 2, transcriptome; Homo sapiens; RNA-Seq
4 mm punch biopsy specimens were incubated in 1% Dispase II (Sigma-Aldrich) in RPMI (Gibco) with 10% FBS for 45 minutes at 37°C, followed by mincing with scalpel, incubation in 0.05% Liberase TL (Sigma Aldrich) in RPMI with 10% fetal bovine serum and 0.008% DNase I (Thermo) for 45 minutes at 37°C, and straining through 40μM mesh. If cryopreservation of sample was necessary (one patient sample), the cell pellet was resuspended in chilled RPMI with 40% FBS, to which was added an equivalent volume of chilled RPMI containing 40% FBS and 30% DMSO. Cells were stored overnight at -80°C. To thaw, the cryovial was placed in a water bath at 37°C for 2m, after which cells were serially diluted 1:1 with warmed RPMI with 10% FBS for a total of 5 dilutions. Viability and cell concentration was assessed using Trypan blue and a hemocytometer. Cells were pelleted and resuspended in phosphate buffered saline with 0.04% bovine serum albumin. From single cell suspensions, libraries were constructed by the Yale Center for Genome Analysis using the Chromium Controller (10X Genomics) with 5' chemistry v2 and V(D)J amplification according to manufacturer's instructions. Libraries were sequenced on an Illumina NovaSeq 6000.