library_construction_protocol: Nuclei were isolated from frozen human brain tumor and non-tumor tissues. Briefly, frozen human tissue samples were homogenized and lysed with Triton X-100 in RNase-free water for nuclei isolation. The isolated nuclei were purified, centrifuged, and resuspended in PBS with BSA and RNAse Inhibitor. The nuclei were diluted to 700 nuclei/ul and loaded to 10x Genomics Chromium Controller to encapsulate single nuclei into droplet emulsions following the manufacturer's recommendations. 3' single nucleus gene expression libraries (Next GEM v3.1, dual index) were constructed using the 10x Genomics Chromium system. Amplified cDNAs and the libraries were measured by Qubit dsDNA HS assay and quality assessed by BioAnalyzer. Each library was sequenced on Illumina NovaSeq 6000 (PE150). Reads were subsequently processed using 10x Genomics Cell Ranger analytical pipeline and human GRCh38 reference genome.