Fig. 6: iTracer-perturb allows for simultaneous genetic perturbation and lineage tracing in mosaic organoids.

a, Schematic for TSC2-targeting or control iTracer-perturb vectors. This system was applied to TSC2 in mosaic cerebral organoids. b, UMAP embedding of scRNA-seq data from 1,673 RFP⁺, 4,766 BFP⁺ and 7,992 GFP⁺ cells sorted from one mosaic iTracer-perturb organoid at 1 month. Cells are colored by cell clusters (bottom) or fluorescent signals (top). c, Feature plot shows marker gene expression. d, Volcano plot shows overall differential expression analysis between RFP⁺ (TSC2-targeting) and BFP⁺ (control) cells, resulting in 385 DEGs (red), 197 of which are unique to the RFP⁺ versus BFP⁺ comparison (dark red). FC, fold change; DE, differentially expressed. e, DAVID functional enrichment analysis of DEGs with significantly increased expression that was uniquely observed in RFP⁺ cells. Bars show −log₁₀ (P) values from tests, and numbers show the number of DEGs annotated with each term. f, UMAP with cells colored by their scar families (≥20 members) and fluorescent signals or by pseudotime (inset). g, Distribution of cells from different scar families (≥20 members) across the constructed pseudotime course.