Fluorescentin vivoediting reporter (FIVER): A novel multispectral reporter ofin vivogenome editing

Advances in genome editing technologies have created opportunities to treat rare genetic diseases, which are often overlooked in terms of therapeutic development. Nonetheless, substantial challenges remain: namely, achieving therapeutically beneficial levels and kinds of editing in the right cell type(s). Here we describe the development of FIVER (fluorescentin vivoediting reporter) — a modular toolkit forin vivodetection of genome editing with distinct fluorescent read-outs for non-homologous end-joining (NHEJ), homology-directed repair (HDR) and homology-independent targeted integration (HITI). We demonstrate that fluorescent outcomes reliably report genetic changes following editing with diverse genome editors in primary cells, organoids andin vivo. We show the potential of FIVER for high-throughput unbiased screens, from small molecule modulators of genome editing outcomes in primary cells through to genome-widein vivoCRISPR cancer screens. Importantly, we demonstrate itsin vivoapplication in postnatal organ systems of interest for genetic therapies — retina and liver. FIVER will broadly help expedite the development of therapeutic genome surgery for many genetic disorders.