CRISPR系统在枯草芽孢杆菌基因编辑中的研究进展

Research advances in CRISPR-based genetic editing of Bacillus subtilis

摘要:

枯草芽孢杆菌( Bacillus subtilis)是一种公认安全的(generally recognized as safe, GRAS)益生菌，同时也是优良的工业生产底盘菌株，具有异源蛋白分泌能力强、低品质碳源生长特性良好以及无明显密码子偏好性等优点。自2016年以来，成簇规律间隔短回文重复序列(clustered regularly interspaced short palindromic repeats, CRISPR)基因编辑技术已成功应用于枯草芽孢杆菌，实现了精准的基因点突变、基因敲除、外源基因插入、基因表达调控及碱基替换等多种基因工程操作。这些进展极大地推动了枯草芽孢杆菌作为高效微生物细胞工厂的发展，并在农业、医药、食品以及合成生物学等领域展现了广泛的应用潜力。本文系统回顾了CRISPR系统在枯草芽孢杆菌中的发展历程，重点总结了其在高效生产不同产物方面的应用成果。旨在通过CRISPR系统靶向优化枯草芽孢杆菌的代谢通路，为在工业化生产中高效、稳定地合成目标产物提供参考，并为新型基因编辑系统的进一步开发与应用提供启示。

Abstract:

Bacillus subtilis is a generally recognized as safe (GRAS) probiotic and an excellent industrial chassis strain. It possesses advantages such as strong heterologous protein secretion capability, robust growth with low-quality carbon sources, and negligible codon bias. Since 2016, clustered regularly interspaced short palindromic repeats (CRISPR)-based gene editing has been successfully applied to B. subtilis, enabling precise genetic modifications, including point mutations, gene knockout, foreign gene insertion, gene expression regulation, and base editing. These advancements have significantly promoted the development of B. subtilis as an efficient microbial cell factory and have shown broad application potential in agriculture, pharmaceuticals, food production, and synthetic biology. This paper systematically review the development of the CRISPR system in B. subtilis and summarize its application in the efficient production of various products. The aim is to provide insights into the targeted optimization of metabolic pathways in B. subtilis via the CRISPR system to achieve efficient and stable industrial production of target products, as well as to offer references for the further development and application of novel gene editing systems.