a tSNE of location-averaged transcriptome for all tumor cells colored by patient. Cancer cells cluster by patient, whereas normal cells from all patients cluster together (encircled clusters indicated by arrows). GSC corresponds to glioma stem cell samples, W corresponds to whole samples. b Enriched glioblastoma stem cell (GSC) gene expression heatmaps showing relative gene expression (raw data) sorted by PC1 per patient. These maps are separated into three rows: top row—100 genes with the lowest value for PC1 loading; bottom row—100 genes with the highest value for PC1 loading; middle row—100 genes with the highest value for PC2 loading. These gene signatures correspond to neuronal, astrocytic, and progenitor signatures, respectively. The TCGA subtype is also shown for each GSC. c Mean and actual rank of genes by PC1 correlation. The actual gene rank (y axis, one point per sample) correlates strongly with the mean gene rank (x axis) in all patients. d Flow cytometry analysis of GSCs and whole-tumor, demonstrating mutually exclusive expression of CD24 and CD44. e Heatmap of gene expression by cNMF signature with associated cell cycle scores and TCGA subtype (right). The most characteristic genes for each signature group are depicted on the x axis. Signatures (y axis) are ordered according to hierarchical clustering (left tree). Left color bar represents the patient sample that generated each signature—patient colors match those in Fig. 1a. Red represents high expression; blue represents low expression. Gene signatures groupings correspond to progenitors, astro-glia (mesenchymal and classical), and neurons, with the addition of cell cycle and hypoxia signatures. cNMF—clustered non-negative matrix factorization. f Heatmap of gene expression by signature ordered by patient as shown by the left color bar. Genes (x axis) are in the same order as Fig. 1e. Patient colors in the color bar match those in Fig. 1a, e. Each patient contains signatures from multiple groups.