Figure 2

Characterisation of single-nuclei G4-CUT&Tag. (a) Schematic diagram of single-nuclei G4 CUT&Tag (snG4-CUT&Tag) workflow. In fixed nuclei, G4-CUT&Tag is used to integrate adapters at G4 sites. Single intact nuclei are then partitioned with barcoded gel beads to index tagmented fragments from individual nuclei using a 10X Genomics Chromium platform with Next GEM Single Cell ATAC Reagents Kits (see “Methods”). (b) Two example G4 landscapes in the human genome generated by G4-CUT&Tag. Blue genomic tracks: normalised read coverages of bulk G4-CUT&Tag libraries (50,000 MCF7 cells and 100,000 U2OS cells); red genomic tracks: normalised signals from aggregated snG4-CUT&Tag libraries; dark red tiles: top 50 single nuclei based on the total number of fragments in the genomic region; pink: Cell Ranger peaks called; black: gene annotations. (c) Clustering of snG4-CUT&Tag data reveals two groups in mixed U2OS and MCF7 sample. t-SNE plot showing dimensionality reduced snG4-CUT&Tag data from mixed cell lines with 671 imputed U2OS (blue) and 467 imputed MCF7 (red) cells. (d) Graphical representation of G4 distribution across promoter G4 peaks in single cells. As in (c) for each t-SNE plot, the top cluster represents individual MCF7 cells while the bottom cluster represents individual U2OS cells. Each dot shows example data covering single U2OS or MCF7 cells for the indicated genes. Grey colouring indicates that no G4 is detected in that cell for Cell Ranger-called promoter peak(s) (1000 bases upstream or 100 bases downstream from the TSS) of the specified genes while orange to red shading quantifies the number of G4s (i.e. pA-Tn5 cut sites) detected in a single cell at the promoter peak(s) of the specified gene. Two example genes (CNGA4 and ALDH6A1) are shown where the G4 promoter peaks are observed in common in the two cell lines, and two example genes (CAPN5 and HMGN1) are shown in which the with G4 promoter peaks specific to only one cell line.