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扩展的pegRNA增强了碱基编辑的编辑能力。

Extended pegRNAs enhance the editing capability of Prime editing.

作者信息

He Kezhang, Xue Qiaomei, Zhou Wei, Wang Pengqi, Hu Xiaodan, Lin Tongtong, Chen Nan, Wang Bowen, Ma Tianhua, Ding Sheng

机构信息

New Cornerstone Science Laboratory, School of Pharmaceutical Sciences, Tsinghua University, Beijing 100084, China.

New Cornerstone Science Laboratory, School of Pharmaceutical Sciences, Tsinghua University, Beijing 100084, China.

出版信息

Trends Biotechnol. 2025 Jan;43(1):206-219. doi: 10.1016/j.tibtech.2024.09.004. Epub 2024 Sep 27.


DOI:10.1016/j.tibtech.2024.09.004
PMID:39341743
Abstract

Genome editing is highly valuable in biomedical research. Despite their versatility, current Prime editing (PE) techniques are limited to short sequence alterations [up to ~44 base pairs (bp)], and exhibit inconsistent or low efficiency across genomic loci, particularly when faced with poly-T sequences. To address these challenges, we developed an extended PE (exPE) technique that can potentially execute any precise genome editing. By harnessing RNA polymerase II (Pol II) promoters to transcribe extended PE guide RNAs (expegRNAs), exPE substantially improves editing efficiency and overcomes the challenges posed by poly-T sequences. Compared with conventional PE, exPE achieves up to a 14-fold increase in the efficiency of base conversions and short insertions, and, remarkably, up to a 259-fold improvement in regions with poly-T sequences. Uniquely, exPE enables seamless insertion of gene-sized DNA fragments into genomes, potentially correcting nearly 90% of human genetic variants, thereby broadening its applications in genetic research and therapy.

摘要

基因组编辑在生物医学研究中具有极高的价值。尽管当前的先导编辑(PE)技术具有多功能性,但它们仅限于短序列改变[最多约44个碱基对(bp)],并且在整个基因组位点上表现出不一致或低效的情况,尤其是在面对多聚T序列时。为应对这些挑战,我们开发了一种扩展先导编辑(exPE)技术,它有可能执行任何精确的基因组编辑。通过利用RNA聚合酶II(Pol II)启动子转录扩展的PE引导RNA(expegRNAs),exPE显著提高了编辑效率,并克服了多聚T序列带来的挑战。与传统的PE相比,exPE在碱基转换和短插入效率上提高了多达14倍,而且在具有多聚T序列的区域中显著提高了多达259倍。独特的是,exPE能够将基因大小的DNA片段无缝插入基因组,有可能校正近90%的人类遗传变异,从而拓宽了其在基因研究和治疗中的应用。

相似文献

[1]
Extended pegRNAs enhance the editing capability of Prime editing.

Trends Biotechnol. 2025-1

[2]
Broadening prime editing toolkits using RNA-Pol-II-driven engineered pegRNA.

Mol Ther. 2022-9-7

[3]
Precise genomic deletions using paired prime editing.

Nat Biotechnol. 2022-2

[4]
Improving prime editing with an endogenous small RNA-binding protein.

Nature. 2024-4

[5]
Deletion and replacement of long genomic sequences using prime editing.

Nat Biotechnol. 2022-2

[6]
A Single Transcript CRISPR-Cas9 System for Multiplex Genome Editing in Plants.

Methods Mol Biol. 2019

[7]
Prime Editing Guide RNA Design Automation Using PINE-CONE.

ACS Synth Biol. 2021-2-19

[8]
High-efficiency prime editing with optimized, paired pegRNAs in plants.

Nat Biotechnol. 2021-8

[9]
WT-PE: Prime editing with nuclease wild-type Cas9 enables versatile large-scale genome editing.

Signal Transduct Target Ther. 2022-4-20

[10]
Highly efficient generation of isogenic pluripotent stem cell models using prime editing.

Elife. 2022-9-7

引用本文的文献

[1]
Enhancing Specificity, Precision, Accessibility, Flexibility, and Safety to Overcome Traditional CRISPR/Cas Editing Challenges and Shape Future Innovations.

Adv Sci (Weinh). 2025-7

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