MAGIC: Mosaic Analysis by gRNA-Induced Crossing-over

Mosaic animals have provided the platform for many fundamental discoveries in developmental biology, cell biology, and other fields. Techniques to produce mosaic animals by mitotic recombination have been extensively developed inDrosophila melanogasterbut are less common for other laboratory organisms. Here, we reportmosaicanalysis bygRNA-inducedcrossing-over (MAGIC), a new technique for generating mosaic animals based on DNA double-strand breaks produced by CRISPR/Cas9. MAGIC efficiently produces mosaic clones in both somatic tissues and the germline ofDrosophila. Further, by developing a MAGIC toolkit for one chromosome arm, we demonstrate the method’s application in characterizing gene function in neural development and in generating fluorescently marked clones in wild-derivedDrosophilastrains. Eliminating the need to introduce recombinase-recognition sites in the genome, this simple and versatile system simplifies mosaic analysis inDrosophilaand can be applied in any organism that is compatible with CRISPR/Cas9.