From cell atlases to mechanisms: bridging scRNA-seq discovery with in vivo genetics

Single-cell RNA sequencing has uncovered vast cellular heterogeneity, yet the in vivo functions of most newly identified subpopulations remain unclear, risking an “atlasing valley of death”, in which descriptive catalogs remain untethered to mechanism. To address this gap, I propose a genetic Sequential Intersectional Recombinase Approach (SIRA) that pairs a cell-type-specific Dre driver with a subpopulation-enriched gene-driven rox-Stop-rox-CreER allele. SIRA restricts Cre activity to cells at the intersection of two genetic markers, allowing precise lineage tracing, targeted ablation, fate manipulation, and floxed-allele perturbation in defined subpopulations. By operationalizing intersectional genetics downstream of discovery-based omics, SIRA provides a practical path from atlas to mechanism in vivo. By the intersection of two markers, SIRA provides a practical route from single-cell atlas data to causal in vivo functional interrogation of cellular subpopulations.