iFlpMosaics enable the multispectral barcoding and high-throughput comparative analysis of mutant and wildtype cells

Abstract To understand gene function, it is necessary to compare cells carrying the mutated target gene with normal cells. In most biomedical studies, the cells being compared are in different mutant and control animals and therefore do not experience the same epigenetic changes and tissue microenvironment. The experimental induction of genetic mosaics is essential to determine a gene cell-autonomous function and to model the etiology of diseases caused by somatic mutations. Current technologies used to induce genetic mosaics in mice lack either accuracy, throughput or barcoding diversity. Here, we present a large set of new genetic tools and mouse lines that enable Flp recombinase-dependent ratiometric induction and single-cell clonal tracking of multiple fluorescently labeled wildtype and Cre-mutant cells within the same time window and tissue microenvironment. The labeled cells can be profiled by multispectral imaging or by FACS and scRNA-seq. This technology facilitates the induction and analysis of genetic mosaics in any cell type and for any given single or combination of floxed genes. iFlpMosaics enables a more accurate understanding of how induced genetic mutations affect the biology of single cells during tissue development, homeostasis, and disease.