title: scATAC-seq of MSK-101 CD45 Sorted IDH-WT Glioma Sample
design: For the scATAC experiment, we followed 10x single-cell ATACseq protocol from 10x Genomics Chromium (Single Cell ATAC Reagent Kits User Guide). Briefly, cells were centrifuged (300g, 5 min, 4 C) and permeabilized with 100 l chilled lysis buffer (10 mM Tris-HCl pH 7.4, 10 mM NaCl, 3 mM MgCl2, 0.1% Tween-20, 0.1% IGEPAL-CA630, 0.01% digitonin and 1% BSA). The samples were incubated on ice for 3-5 min and resuspended with 1 ml chilled wash buffer (10 mM Tris-HCl pH 7.4, 10 mM NaCl, 3 mM MgCl2, 0.1% Tween-20 and 1% BSA). After centrifugation (500g, 5 min, 4 C), the pellets were resuspended in 100 l chilled Nuclei buffer (10x Genomics). Nuclei were counted using a haemocytometer, and finally the nucleus concentration was adjusted to 3,000 nuclei per l. We used 15,360 nuclei as input for tagmentation. Nuclei were diluted to 5 l with 1 nuclei buffer (10x Genomics) and mixed with ATAC buffer (10x Genomics) and ATAC enzyme (10x Genomics) for tagmentation (60 min, 37 C). Single-cell ATACseq libraries were generated using the Chromium Chip E Single Cell ATAC kit (10x Genomics, 1000086) and indexes (Chromium i7 Multiplex Kit N, Set A, 10x Genomics, 1000084) following the manufacturers instructions. Final libraries were quantified using a Qubit fluorimeter (Life Technologies) and the nucleosomal pattern was verified using a BioAnalyzer (Agilent). Libraries were sequenced on a NovaqSeq6000 (Illumina) with the following read lengths: 50 + 8 + 16 + 50 (Read1 + Index1 + Index2 + Read2).