[논문]A versatile genetic engineering toolkit for E. coli based on CRISPR-prime editing

Tong, Yaojun; Jørgensen, Tue S.; Whitford, Christopher M.; Weber, Tilmann; Lee, Sang Yup

doi:10.1038/s41467-021-25541-3

[해외논문] A versatile genetic engineering toolkit for E. coli based on CRISPR-prime editing 원문보기

Nature communications, v.12 no.1, 2021년, pp.5206 -

Tong, Yaojun (The Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Lyngby, Denmark) , Jørgensen, Tue S. (The Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Lyngby, Denmark) , Whitford, Christopher M. (The Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Lyngby, Denmark) , Weber, Tilmann (The Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Lyngby, Denmark) , Lee, Sang Yup (The Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Lyngby, Denmark)

Abstract ▼ AI-Helper

CRISPR base editing is a powerful method to engineer bacterial genomes. However, it restricts editing to single-nucleotide substitutions. Here, to address this challenge, we adapt a CRISPR-Prime Editing-based, DSB-free, versatile, and single-nucleotide resolution genetic manipulation toolkit for prokaryotes. It can introduce substitutions, deletions, insertions, and the combination thereof, both in plasmids and the chromosome of E. coli with high fidelity. Notably, under optimal conditions, the efficiency of 1-bp deletions reach up to 40%. Moreover, deletions of up to 97 bp and insertions up to 33 bp were successful with the toolkit in E. coli, however, efficiencies dropped sharply with increased fragment sizes. With a second guide RNA, our toolkit can achieve multiplexed editing albeit with low efficiency. Here we report not only a useful addition to the genome engineering arsenal for E. coli, but also a potential basis for the development of similar toolkits for other bacteria.CRISPR prime editing enables double-strand break free engineering of the genome. Here the authors present a toolkit for prime editing in E. coli.

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용어 설명: PCR은 세균 특이성이 있는 primer를 이용하여 적은 수의 세균이 있을지라도 쉽게 검출할 수 있는 유용한 방법이며, 이를 이용하여 구강 내 치면세균막이나 타액에서 직접 세균을 검출할 수 있게 되었다[8].

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