Genome editing is induced in a binary manner in single human cells.
Genome editing is induced in a binary manner in single human cells.
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DOI：
10.1016/j.isci.2022.105619
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发表时间：
2022-12-22
期刊：
影响因子：
5.8
通讯作者：
Miyaoka, Yuichiro
中科院分区：
文献类型：
--
作者：
Takahashi, Gou;Kondo, Daiki;Maeda, Minato;Morishita, Yuji;Miyaoka, Yuichiro
即使打算进行精确的核苷酸操作，基因组编辑的结果也可能是不同的，通常包括随机插入和删除。这些不同结果在单个细胞中的组合和频率不仅在转基因细胞系的产生中至关重要，而且在评估基因组编辑疗法的临床效果方面也是至关重要的。然而，目前的方法只分析细胞群体，而不是单个细胞。在这里，我们利用单粒子分离系统(SPIS)有效地分离单个细胞，系统地分析单个人类培养细胞的基因组编辑结果。结果，我们发现基因组编辑诱导具有二元性，即细胞的目标等位基因往往都被编辑或根本不编辑。这项研究加深了我们对基因组编辑诱导模式的理解，并为分析单细胞基因组编辑结果提供了一种新的策略。使用Spis Cas9分离了2,600多个经基因组编辑的人类细胞克隆，在单细胞中，HDR要么完全编辑目标等位基因，要么没有编辑。在单细胞生物科学；生物技术；生物技术；遗传工程中，HDR更常伴随NHEJ而不是WT。
Even when precise nucleotide manipulations are intended, the outcomes of genome editing can be diverse, often including random insertions and deletions. The combinations and frequencies of these different outcomes in single cells are critical not only in the generation of genetically modified cell lines but also in the evaluation of the clinical effects of genome editing therapies. However, current methods only analyze cell populations, not single cells. Here, we utilized the Single Particle isolation System (SPiS) for the efficient isolation of single cells to systematically analyze genome editing results in individual human cultured cells. As a result, we discovered that genome editing induction has a binary nature, that is, the target alleles of cells tend to be all edited or not edited at all. This study enhances our understanding of the induction pattern of genome editing and provides a new strategy to analyze genome editing outcomes in single cells. Over 2,600 genome-edited human cell clones were isolated by using SPiS Cas9 exhibited either full editing of target alleles or no editing in single cells HDR was more often accompanied by NHEJ than WT in single cells Biological sciences; Bioengineering; Biotechnology; Genetic engineering.
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