Rxivist combines preprints from bioRxiv with data from Twitter to help you find the papers being discussed in your field. Currently indexing 70,260 bioRxiv papers from 306,813 authors.
No unexpected CRISPR-Cas9 off-target activity revealed by trio sequencing of gene-edited mice
CRISPR-Cas technologies have transformed genome-editing of experimental organisms and have immense therapeutic potential. Despite significant advances in our understanding of the CRISPR-Cas9 system, concerns remain over the potential for off-target effects. Recent studies have addressed these concerns using whole-genome sequencing (WGS) of gene-edited embryos or animals to search for de novo mutations (DNMs), which may represent candidate changes induced by poor editing fidelity. Critically, these studies used strain-matched but not pedigree-matched controls and thus were unable to reliably distinguish generational or colony-related differences from true DNMs. Here we used a trio design and whole genome sequenced 8 parents and 19 embryos, where 10 of the embryos were mutagenised with well-characterised gRNAs targeting the coat colour Tyrosinase (Tyr) locus. Detailed analyses of these whole genome data allowed us to conclude that if CRISPR mutagenesis were causing SNV or indel off-target mutations in treated embryos, then the number of these mutations is not statistically distinguishable from the background rate of DNMs occurring due to other processes.
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