Bibliographic Details
| Title: |
High-efficiency targeted transgene integration via primed micro-homologues |
| Authors: |
Chenxin Wang, Sen Fang, Yangcan Chen, Na Tang, Guanyi Jiao, Yanping Hu, Jing Li, Qingtong Shan, Xin Wang, Guihai Feng, Qi Zhou, Wei Li |
| Source: |
Cell Discovery, Vol 9, Iss 1, Pp 1-14 (2023) |
| Publisher Information: |
Nature Publishing Group, 2023. |
| Publication Year: |
2023 |
| Collection: |
LCC:Cytology |
| Subject Terms: |
Cytology, QH573-671 |
| More Details: |
Abstract Due to the difficulties in precisely manipulating DNA repair pathways, high-fidelity targeted integration of large transgenes triggered by double-strand breaks is inherently inefficient. Here, we exploit prime editors to devise a robust knock-in (KI) strategy named primed micro-homologues-assisted integration (PAINT), which utilizes reverse-transcribed single-stranded micro-homologues to boost targeted KIs in different types of cells. The improved version of PAINT, designated PAINT 3.0, maximizes editing efficiency and minimizes off-target integration, especially in dealing with scarless in-frame KIs. Using PAINT 3.0, we target a reporter transgene into housekeeping genes with editing efficiencies up to 80%, more than 10-fold higher than the traditional homology-directed repair method. Moreover, the use of PAINT 3.0 to insert a 2.5-kb transgene achieves up to 85% KI frequency at several therapeutically relevant genomic loci, suggesting its potential for clinical applications. Finally, PAINT 3.0 enables high-efficiency non-viral genome targeting in primary T cells and produces functional CAR-T cells with specific tumor-killing ability. Thus, we establish that the PAINT method is a powerful gene editing tool for large transgene integrations and may open new avenues for cell and gene therapies and genome writing technologies. |
| Document Type: |
article |
| File Description: |
electronic resource |
| Language: |
English |
| ISSN: |
2056-5968 |
| Relation: |
https://doaj.org/toc/2056-5968 |
| DOI: |
10.1038/s41421-023-00552-0 |
| Access URL: |
https://doaj.org/article/ca7c59d5d76a47a0a26822f896419b21 |
| Accession Number: |
edsdoj.7c59d5d76a47a0a26822f896419b21 |
| Database: |
Directory of Open Access Journals |
