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Background <p><i>Vibrio</i> 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.</p> Results <p>The CRISPR-based cytosine base editing (CBE) system was established in <i>Vibrio</i> 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.</p> Conclusions <p>Developing a highly efficient CBE system and improving transformation efficiency enable versatile genetic manipulation of <i>Vibrio</i> sp. dhg for diverse engineering in brown macroalgae bioconversion.</p>

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

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2025年4月9日

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

摘要

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.