Development of a CRISPR-based cytosine base editor for restriction-modification system inactivation to enhance transformation efficiency in Vibrio Sp. dhg.

IF 5.7 3区生物学 Q1 BIOCHEMICAL RESEARCH METHODS

Journal of Biological Engineering Pub Date : 2025-04-09 DOI:10.1186/s13036-025-00500-4

Yang Jun Shon, Dongyeop Baek, Su Bin Jin, Woo Jae Kim, Gyoo Yeol Jung, Hyun Gyu Lim

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引用次数: 0

Abstract

Background: Vibrio sp. dhg is a fast-growing, alginate-utilizing, marine bacterium being developed as a platform host for macroalgae biorefinery. To maximize its potential in the production of various value-added products, there is a need to expand genetic engineering tools for versatile editing.

Results: The CRISPR-based cytosine base editing (CBE) system was established in Vibrio sp. dhg, enabling C: G-to-T: A point mutations in multiple genomic loci. This CBE system displayed high editing efficiencies for single and multiple targets, reaching up to 100%. The CBE system efficiently introduced premature stop codons, inactivating seven genes encoding putative restriction enzymes of the restriction-modification system in two rounds. A resulting engineered strain displayed significantly enhanced transformation efficiency by up to 55.5-fold.

Conclusions: Developing a highly efficient CBE system and improving transformation efficiency enable versatile genetic manipulation of Vibrio sp. dhg for diverse engineering in brown macroalgae bioconversion.

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基于crispr的胞嘧啶碱基编辑器的开发，用于限制修饰系统失活以提高弧菌的转化效率。

背景：弧菌（Vibrio sp. dhg）是一种快速生长、利用海藻酸盐的海洋细菌，正被开发为大型藻类生物炼制的平台宿主。为了最大限度地发挥其在生产各种增值产品方面的潜力，有必要扩大用于多功能编辑的基因工程工具。结果：在Vibrio sp. dhg中建立了基于crispr的胞嘧啶碱基编辑（CBE）系统，实现了多个基因组位点的C: g到t: A点突变。该CBE系统显示出较高的单目标和多目标编辑效率，最高可达100%。CBE系统有效地引入过早终止密码子，分两轮灭活了7个编码限制性内切酶的基因。由此产生的工程菌株显示出显著提高的转化效率高达55.5倍。结论：建立高效的CBE系统，提高转化效率，为实现大型褐藻生物转化工程中弧菌（Vibrio sp. dhg）的多样化基因操作提供了可能。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Journal of Biological Engineering BIOCHEMICAL RESEARCH METHODS-BIOTECHNOLOGY & APPLIED MICROBIOLOGY

CiteScore

7.10

自引率

1.80%

发文量

审稿时长

17 weeks

期刊介绍： Biological engineering is an emerging discipline that encompasses engineering theory and practice connected to and derived from the science of biology, just as mechanical engineering and electrical engineering are rooted in physics and chemical engineering in chemistry. Topical areas include, but are not limited to: Synthetic biology and cellular design Biomolecular, cellular and tissue engineering Bioproduction and metabolic engineering Biosensors Ecological and environmental engineering Biological engineering education and the biodesign process As the official journal of the Institute of Biological Engineering, Journal of Biological Engineering provides a home for the continuum from biological information science, molecules and cells, product formation, wastes and remediation, and educational advances in curriculum content and pedagogy at the undergraduate and graduate-levels. Manuscripts should explore commonalities with other fields of application by providing some discussion of the broader context of the work and how it connects to other areas within the field.