Abstract：
The type I CRISPR system has recently emerged as a promising tool, especially for large-scale genomic modification, but its application to generate model animals by editing zygotes had not been established. In this study, we demonstrate genome editing in zygotes using the type I-E CRISPR-Cas3 system, which efficiently generates deletions of several thousand base pairs at targeted loci in mice with 40%-70% editing efficiency without off-target mutations. To overcome the difficulties associated with detecting the variable deletions, we used a newly long-read sequencing-based multiplex genotyping approach. Demonstrating remarkable versatility, our Cas3-based technique was successfully extended to rats as well as mice, even by zygote electroporation methods. Knockin for SNP exchange and genomic replacement with a donor plasmid were also achieved in mice. This pioneering work with the type I CRISPR zygote editing system offers increased flexibility and broader applications in genetic engineering across different species.
摘要：
I型CRISPR系统最近成为一种有前途的工具，特别是对于大规模的基因组改造，但是它在通过编辑受精卵产生模型动物中的应用尚未建立。在这项研究中,我们证明了使用I-E型CRISPR-Cas3系统在受精卵中进行基因组编辑，在没有脱靶突变的情况下，其在小鼠中以40％-70％的编辑效率有效地在目标基因座处产生数千个碱基对的缺失。为了克服与检测变量缺失相关的困难，我们使用了一种新的基于长读数测序的多重基因分型方法.展示了非凡的多功能性，我们基于Cas3的技术被成功地扩展到大鼠和小鼠，甚至通过合子电穿孔方法。在小鼠中也实现了SNP交换和用供体质粒进行基因组置换的敲入。I型CRISPR受精卵编辑系统的这项开创性工作在不同物种的基因工程中提供了更大的灵活性和更广泛的应用。
