GSM5155454: LNCaP RES-B [scATAC-seq]; Homo sapiens; ATAC-seq
Cell nuclei were isolated following the 10x Genomics Demonstrated Protocol for Single Cell ATAC Sequencing (CG000169-Rev C). Briefly, the cell suspension was washed once in PBS/0.04%BSA, and 2 x 105 cells were pelleted (300 x g for 5 min), resuspended in 100 µl freshly prepared Lysis Buffer (10 mM Tris-HCl pH 7.4, 10 mM NaCl, 3 mM MgCl2, 0.1% Tween-20, 0.1% NP40 Substitute, 0.01% Digitonin, 1% BSA), and incubated on ice for 4 min (LNCaP parental cells), 6 min (RES-A), or 5 min (RES-B). The lysates were diluted with 1 ml wash buffer (10 mM Tris-HCl pH 7.4, 10 mM NaCl, 3 mM MgCl2, 0.1% Tween-20, 1% BSA), and the nuclei were pelleted (500 x g for 5 min) and resuspended in 30 µl 1x Nuclei Buffer (10x Genomics PN-2000153). Successful preparation of intact, isolated nuclei was confirmed through visual inspection in a phase-contrast microscopy, and nuclei concentration was assessed with a Countess II FL Automated Cell Counter Single Cell ATAC sequencing using 10x Chromium, 10x Genomics library preparation and the Chromium Single Cell ATAC Reagent Kits (v1) User Guide (CG000168 Rev D).