Versatile and efficient mammalian genome editing with Type I-C CRISPR System of Desulfovibrio vulgaris.

IF 8 2区 生物学 Q1 BIOLOGY
Science China Life Sciences Pub Date : 2024-11-01 Epub Date: 2024-08-07 DOI:10.1007/s11427-023-2682-5
Pan Li, Dingcai Dong, Fei Gao, Yuyang Xie, Honglin Huang, Siwei Sun, Zhao Ma, Cheng He, Jinsheng Lai, Xuguang Du, Sen Wu
{"title":"Versatile and efficient mammalian genome editing with Type I-C CRISPR System of Desulfovibrio vulgaris.","authors":"Pan Li, Dingcai Dong, Fei Gao, Yuyang Xie, Honglin Huang, Siwei Sun, Zhao Ma, Cheng He, Jinsheng Lai, Xuguang Du, Sen Wu","doi":"10.1007/s11427-023-2682-5","DOIUrl":null,"url":null,"abstract":"<p><p>CRISPR-Cas tools for mammalian genome editing typically rely on single Cas9 or Cas12a proteins. While type I CRISPR systems in Class I may offer greater specificity and versatility, they are not well-developed for genome editing. Here, we present an alternative type I-C CRISPR system from Desulfovibrio vulgaris (Dvu) for efficient and precise genome editing in mammalian cells and animals. We optimized the Dvu type I-C editing complex to generate precise deletions at multiple loci in various cell lines and pig primary fibroblast cells using a paired PAM-in crRNA strategy. These edited pig cells can serve as donors for generating transgenic cloned piglets. The Dvu type I-C editor also enabled precise large fragment replacements with homology-directed repair. Additionally, we adapted the Dvu-Cascade effector for cytosine and adenine base editing, developing Dvu-CBE and Dvu-ABE systems. These systems efficiently induced C-to-T and A-to-G substitutions in human genes without double-strand breaks. Off-target analysis confirmed the high specificity of the Dvu type I-C editor. Our findings demonstrate the Dvu type I-C editor's potential for diverse mammalian genome editing applications, including deletions, fragment replacement, and base editing, with high efficiency and specificity for biomedicine and agriculture.</p>","PeriodicalId":21576,"journal":{"name":"Science China Life Sciences","volume":null,"pages":null},"PeriodicalIF":8.0000,"publicationDate":"2024-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Science China Life Sciences","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1007/s11427-023-2682-5","RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2024/8/7 0:00:00","PubModel":"Epub","JCR":"Q1","JCRName":"BIOLOGY","Score":null,"Total":0}
引用次数: 0

Abstract

CRISPR-Cas tools for mammalian genome editing typically rely on single Cas9 or Cas12a proteins. While type I CRISPR systems in Class I may offer greater specificity and versatility, they are not well-developed for genome editing. Here, we present an alternative type I-C CRISPR system from Desulfovibrio vulgaris (Dvu) for efficient and precise genome editing in mammalian cells and animals. We optimized the Dvu type I-C editing complex to generate precise deletions at multiple loci in various cell lines and pig primary fibroblast cells using a paired PAM-in crRNA strategy. These edited pig cells can serve as donors for generating transgenic cloned piglets. The Dvu type I-C editor also enabled precise large fragment replacements with homology-directed repair. Additionally, we adapted the Dvu-Cascade effector for cytosine and adenine base editing, developing Dvu-CBE and Dvu-ABE systems. These systems efficiently induced C-to-T and A-to-G substitutions in human genes without double-strand breaks. Off-target analysis confirmed the high specificity of the Dvu type I-C editor. Our findings demonstrate the Dvu type I-C editor's potential for diverse mammalian genome editing applications, including deletions, fragment replacement, and base editing, with high efficiency and specificity for biomedicine and agriculture.

利用普通脱硫弧菌的 I-C 型 CRISPR 系统进行多功能、高效的哺乳动物基因组编辑。
用于哺乳动物基因组编辑的 CRISPR-Cas 工具通常依赖单个 Cas9 或 Cas12a 蛋白。虽然I类CRISPR系统可能具有更高的特异性和多功能性,但它们在基因组编辑方面并不发达。在这里,我们介绍了一种来自Desulfovibrio vulgaris(Dvu)的I-C型CRISPR系统,用于在哺乳动物细胞和动物中进行高效、精确的基因组编辑。我们对 Dvu I-C 型编辑复合物进行了优化,利用成对的 PAM-in crRNA 策略在各种细胞系和猪原代成纤维细胞中生成多个位点的精确缺失。这些经过编辑的猪细胞可作为供体,用于产生转基因克隆仔猪。Dvu I-C 型编辑器还能通过同源定向修复实现大片段的精确替换。此外,我们还将 Dvu 级联效应器用于胞嘧啶和腺嘌呤碱基编辑,开发出了 Dvu-CBE 和 Dvu-ABE 系统。这些系统能有效地诱导人类基因中的 C 到 T 和 A 到 G 的置换,而不会发生双链断裂。脱靶分析证实了 Dvu I-C 型编辑器的高度特异性。我们的研究结果表明,Dvu I-C 型编辑器可用于多种哺乳动物基因组编辑应用,包括缺失、片段置换和碱基编辑,在生物医学和农业领域具有高效率和特异性。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
求助全文
约1分钟内获得全文 求助全文
来源期刊
CiteScore
15.10
自引率
8.80%
发文量
2907
审稿时长
3.2 months
期刊介绍: Science China Life Sciences is a scholarly journal co-sponsored by the Chinese Academy of Sciences and the National Natural Science Foundation of China, and it is published by Science China Press. The journal is dedicated to publishing high-quality, original research findings in both basic and applied life science research.
×
引用
GB/T 7714-2015
复制
MLA
复制
APA
复制
导出至
BibTeX EndNote RefMan NoteFirst NoteExpress
×
提示
您的信息不完整,为了账户安全,请先补充。
现在去补充
×
提示
您因"违规操作"
具体请查看互助需知
我知道了
×
提示
确定
请完成安全验证×
copy
已复制链接
快去分享给好友吧!
我知道了
右上角分享
点击右上角分享
0
联系我们:info@booksci.cn Book学术提供免费学术资源搜索服务,方便国内外学者检索中英文文献。致力于提供最便捷和优质的服务体验。 Copyright © 2023 布克学术 All rights reserved.
京ICP备2023020795号-1
ghs 京公网安备 11010802042870号
Book学术文献互助
Book学术文献互助群
群 号:481959085
Book学术官方微信