CRISPR-Cas9基因组编辑在遗传病和癌症的治疗潜力上有着很大的希望，但其安全性也是一个关注点。在本研究中，我们使用10x联合读取测序和光学基因组映射来对编辑后的基因组完整性进行全基因组分析，并将其与四个亲本细胞系进行比较。除了之前报道的位点上的大结构变异外，我们还在两个编辑的细胞系中发现了以往意外的非同源非靶位点上两个大染色体缺失（91.2和136Kb），这些缺失与sgRNA序列没有相似性。CRISPR-Cas9编辑引起的观察到的大结构变异可能在分裂细胞中导致病理性后果，从而限制了CRISPR-Cas9编辑系统在疾病建模和基因治疗中的实用性。在这项研究中，我们的全基因组分析可能提供了一个宝贵的策略，以确保基因组编辑后的基因组完整性，以减少研究和临床应用中意外效应的风险。© 2023 Springer Nature Limited.

CRISPR-Cas9 genome editing has promising therapeutic potential for genetic diseases and cancers, but safety could be a concern. Here we use whole genomic analysis by 10x linked-read sequencing and optical genome mapping to interrogate the genome integrity after editing and in comparison to four parental cell lines. In addition to the previously reported large structural variants at on-target sites, we identify heretofore unexpected large chromosomal deletions (91.2 and 136 Kb) at atypical non-homologous off-target sites without sequence similarity to the sgRNA in two edited lines. The observed large structural variants induced by CRISPR-Cas9 editing in dividing cells may result in pathogenic consequences and thus limit the usefulness of the CRISPR-Cas9 editing system for disease modeling and gene therapy. In this work, our whole genomic analysis may provide a valuable strategy to ensure genome integrity after genomic editing to minimize the risk of unintended effects in research and clinical applications.© 2023. Springer Nature Limited.