A design optimized prime editor with expanded scope and capability in plants
A design optimized prime editor with expanded scope and capability in plants
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DOI：
10.1038/s41477-021-01043-4
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发表时间：
2021-12-23
期刊：
影响因子：
18
通讯作者：
Yang, Jinxiao
中科院分区：
文献类型：
--
作者：
Xu, Wen;Yang, Yongxing;Yang, Jinxiao
以可编程方式操纵基因组的能力照亮了生物学，并在植物育种中显示出前景。 Prime 编辑是一种多功能基因编辑方法，可直接将新的遗传信息写入指定的 DNA 位点，而不需要双链 DNA 断裂，但在植物中效率较低(1-5)。在本研究中，N 端逆转录酶-Cas9 切口酶融合体在水稻中的表现优于常用的 C 端融合体。此外，在逆转录酶模板中引入多核苷酸取代可以提高引物编辑的效率。通过协同使用这两种方法，实现了水稻转基因植物中13个内源靶点的平均编辑频率高达24.3%、玉米原生质体中4个靶点的6.2%和人类细胞中12.5%的prime编辑，比原始编辑器Prime Editor 3高出两到三倍。因此，我们的优化方法有潜力使更多以前不可编辑的靶位点变得可编辑，并扩展了prime的范围和功能。将来进行编辑。
The ability to manipulate the genome in a programmable manner has illuminated biology and shown promise in plant breeding. Prime editing, a versatile gene-editing approach that directly writes new genetic information into a specified DNA site without requiring double-strand DNA breaks, suffers from low efficiency in plants(1-5). In this study, N-terminal reverse transcriptase-Cas9 nickase fusion performed better in rice than the commonly applied C-terminal fusion. In addition, introduction of multiple-nucleotide substitutions in the reverse transcriptase template stimulated prime editing with enhanced efficiency. By using these two methods synergistically, prime editing with an average editing frequency as high as 24.3% at 13 endogenous targets in rice transgenic plants, 6.2% at four targets in maize protoplasts and 12.5% in human cells was achieved, which is two- to threefold higher than the original editor, Prime Editor 3. Therefore, our optimized approach has potential to make more formerly non-editable target sites editable, and expands the scope and capabilities of prime editing in the future.