High-Resolution Lineage Tracing in Native Tissue Contexts [SPACE-seq]

High-resolution lineage tracing has provided essential insights into key biological processes, yet existing methods are limited in tracing diverse cell populations at single-cell resolution while preserving spatial context. Here, we present SPACE-seq (SPatial trAcing enabled by CRISPR-based barcodEs and Slide-seq), a versatile tracing platform that integrates spatial transcriptomics with Cas9 barcoding in mice, to allow simultaneous analysis of cell lineages, transcriptomic states, and spatial organization at near-cellular resolution. Using this approach, we generated lineage maps that reveal novel interactions during liver cancer progression within the tumor microenvironment and uncovered distinct clonal architecture patterns in the developing cerebral cortex. Additionally, we identified a developmentally restricted window for hepatoblast migration and spatially confined compartments involved in liver lobe formation. With its broad applicability and adaptability, SPACE-seq offers new potential to explore previously inaccessible questions in cellular organization and lineage dynamics.