Tools for experimental and computational analyses of off-target editing by programmable nucleases.
Tools for experimental and computational analyses of off-target editing by programmable nucleases.
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DOI：
10.1038/s41596-020-00431-y
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发表时间：
2021-01
期刊：
影响因子：
14.8
通讯作者：
Bao G
中科院分区：
文献类型：
--
作者：
Bao XR;Pan Y;Lee CM;Davis TH;Bao G
Genome editing using programmable nucleases is revolutionizing life science and medicine. Off-target editing by these nucleases remains a considerable concern, especially in therapeutic applications. Here we review tools developed for identifying potential off-target editing sites and compare the ability of these tools to properly analyze off-target effects. Recent advances in both in silico and experimental tools for off-target analysis have generated remarkably concordant results for sites with high off-target editing activity. However, no single tool is able to accurately predict low-frequency off-target editing, presenting a bottleneck in therapeutic genome editing, because even a small number of cells with off-target editing can be detrimental. Therefore, we recommend that at least one in silico tool and one experimental tool should be used together to identify potential off-target sites, and amplicon-based next-generation sequencing (NGS) should be used as the gold standard assay for assessing the true off-target effects at these candidate sites. Future work to improve off-target analysis includes expanding the true off-target editing dataset to evaluate new experimental techniques and to train machine learning algorithms; performing analysis using the particular genome of the cells in question rather than the reference genome; and applying novel NGS techniques to improve the sensitivity of amplicon-based off-target editing quantification.
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