GSM4983569: pCGY4021_recurrent; Homo sapiens; ATAC-seq
Flash frozen tissue samples were resuspended in cold ATAC resuspension buffer (10 mM Tris-HCl pH 7.4, 10 mM NaCl, 3 mM MgCl2, 0.1% NP-40, 0.1% Tween-20, 0.01% Digitonin, 1% BSA in PBS) on ice and dissociated using a wide-bore P1000 pipette tip and vortexing, followed by 10 minutes of incubation on ice. Cells were spun down at 500x g for 5 minutes at 4 C, washed in the ATAC resuspension buffer, spun down again, and resuspended in ATAC-Tween wash buffer (10 mM Tris-HCl pH 7.4, 10 mM NaCl, 3 mM MgCl2, 0.1% Tween-20, 1% BSA in PBS), then passed through a cell strainer top FACS tube (Falcon; #38030) to remove debris. Nuclei quality and quantity was evaluated using trypan blue on an Invitrogen Countess II device in duplicate, and a subset of nuclei was spun down in a fresh tube and resuspended in 10X sample dilution buffer. Libraries were prepared according to 10X Genomics Chromium Single Cell ATAC Library & Gel Bead Kit Chemistry. Briefly, single nuclei were first treated in bulk with Tn5 transposase to preferentially insert sequencing adaptors into accessible DNA regions. Transposed nuclei are then partitioned into GEMs in the 10x Chromium Controller. All the DNA fragments from the same nucleus share a common 10x barcode. The barcoded, accessible DNA fragments are subsequently pooled for downstream processing and library preparation. Completed libraries were further quality checked for fragment size and distribution using an Agilent TapeStation prior to sequencing.Next generation sequencing was performed using Illumina NextSeq 500  per cell. 