IT-scC&T-seq streamlines scalable, parallel profiling of protein-DNA interactions in single cells

Single-cell profiling protein-chromatin interactions is often constrained by complex workflows, high cost, or dependence on specialised equipment. We present indexed tagmentation-based single-cell CUT&Tag-sequencing (IT-scC&T-seq), a modular, plate-based strategy using three-round combinatorial barcoding. IT-scC&T-seq robustly profiles histone modifications and transcription factors with high specificity and throughput, supporting simultaneous analysis of multiple samples and epitopes. Notably, it enables sensitive single-cell mapping of lamina-associated domains, low-abundance chromatin features previously difficult to resolve. Applied to adult mouse mammary gland, the method reveals cell-type-specific chromatin landscapes and lineage-regulatory dynamics. Together, IT-scC&T-seq provides a scalable, cost-effective, and broadly accessible approach for high-resolution chromatin profiling. Indexed tagmentation-based single-cell CUT&Tag-sequencing (IT-scC&T-seq) of (1) H3K4me3 in mixed HEK293T and B6 mouse embyronic stem cells; (2) H3K4me3, H3K27me3, RNA-PolII and CTCF in K562; (3) LMNB1 in mixed H1 human embyronic stem cells, K562, HEK293T, and H1-derived MSCs; (4) multiple epitope (H3K4me3, H3K36me3, H3K27me3 and CTCF) across MSCs, H1-ESCs, and HEK293T; (5) H3K4me3 and H3K27ac in 10-week female WT mice mammary gland.