Cas9/Nickase-induced allelic conversion by homologous chromosome-templated repair in <i>Drosophila</i> somatic cells
Sitara Roy, Sara Sanz Juste, Marketta Sneider, Ankush Auradkar, Carissa F. Klanseck, Zhiqian Li, Alison Henrique Ferreira Julio, Víctor López Del Amo, Ethan Bier, Annabel Guichard
Abstract
Repair of double-strand breaks (DSBs) in somatic cells is primarily accomplished by error-prone nonhomologous end joining and less frequently by precise homology-directed repair preferentially using the sister chromatid as a template. Here, a Drosophila system performs efficient somatic repair of both DSBs and single-strand breaks (SSBs) using intact sequences from the homologous chromosome in a process we refer to as homologous chromosome-templated repair (HTR). Unexpectedly, HTR-mediated allelic conversion at the white locus was more efficient (40 to 65%) in response to SSBs induced by Cas9-derived nickases D10A or H840A than to DSBs induced by fully active Cas9 (20 to 30%). Repair phenotypes elicited by Nickase versus Cas9 differ in both developmental timing (late versus early stages, respectively) and the production of undesired mutagenic events (rare versus frequent). Nickase-mediated HTR represents an efficient and unanticipated mechanism for allelic correction, with far-reaching potential applications in the field of gene editing.