Optimized nickase- and nuclease-based prime editing in human and mouse cells.
Optimized nickase- and nuclease-based prime editing in human and mouse cells.
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DOI：
10.1093/nar/gkab792
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发表时间：
2021-10-11
影响因子：
14.9
通讯作者：
Thomas PQ
中科院分区：
文献类型：
--
作者：
Adikusuma F;Lushington C;Arudkumar J;Godahewa GI;Chey YCJ;Gierus L;Piltz S;Geiger A;Jain Y;Reti D;Wilson LOW;Bauer DC;Thomas PQ
使用引物编辑（PE）的精确基因组修饰在研究和临床应用中具有巨大的潜力。在这项研究中，我们生成了一体化的主要编辑（PEA 1）构建体，该构建体携带PE所需的所有组件，沿着的还有选择标记。我们在HEK293T、K562、HeLa和小鼠胚胎干（ES）细胞中测试了这些构建体（具有选择）。我们发现HEK293T细胞中的PE效率比以前观察到的要高得多，高达95%（平均67%）。然而，在K562和HeLa细胞中的效率仍然很低。为了提高K562和HeLa中的PE效率，我们生成了核酸酶引物编辑器，并在这些细胞系以及小鼠ES细胞中测试了该系统。PE-核酸酶大大增加了引物编辑起始，然而，预期编辑的安装通常伴随着来自修复模板的额外插入。最后，我们表明，合子注射的核酸酶引物编辑器可以产生正确的修改在小鼠胎儿高达100%的效率。
Precise genomic modification using prime editing (PE) holds enormous potential for research and clinical applications. In this study, we generated all-in-one prime editing (PEA1) constructs that carry all the components required for PE, along with a selection marker. We tested these constructs (with selection) in HEK293T, K562, HeLa and mouse embryonic stem (ES) cells. We discovered that PE efficiency in HEK293T cells was much higher than previously observed, reaching up to 95% (mean 67%). The efficiency in K562 and HeLa cells, however, remained low. To improve PE efficiency in K562 and HeLa, we generated a nuclease prime editor and tested this system in these cell lines as well as mouse ES cells. PE-nuclease greatly increased prime editing initiation, however, installation of the intended edits was often accompanied by extra insertions derived from the repair template. Finally, we show that zygotic injection of the nuclease prime editor can generate correct modifications in mouse fetuses with up to 100% efficiency.
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影响因子：
12.3
作者：
Park SJ;Jeong TY;Shin SK;Yoon DE;Lim SY;Kim SP;Choi J;Lee H;Hong JI;Ahn J;Seong JK;Kim K
通讯作者：
Kim K
影响因子：
16.6
作者：
Schene IF;Joore IP;Oka R;Mokry M;van Vugt AHM;van Boxtel R;van der Doef HPJ;van der Laan LJW;Verstegen MMA;van Hasselt PM;Nieuwenhuis EES;Fuchs SA
通讯作者：
Fuchs SA
DOI：
10.1073/pnas.2021996118
发表时间：
2021-01-05
影响因子：
11.1
作者：
Bosch, Justin A.;Birchak, Gabriel;Perrimon, Norbert
通讯作者：
Perrimon, Norbert
DOI：
10.1089/crispr.2020.0022
发表时间：
2021-03
期刊：
The CRISPR journal
影响因子：
--
作者：
Bloh K;Kanchana R;Bialk P;Banas K;Zhang Z;Yoo BC;Kmiec EB
通讯作者：
Kmiec EB
影响因子：
16.6
作者：
Liu P;Liang SQ;Zheng C;Mintzer E;Zhao YG;Ponnienselvan K;Mir A;Sontheimer EJ;Gao G;Flotte TR;Wolfe SA;Xue W
通讯作者：
Xue W