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在Bifidobacterium中高效的CRISPR-Cas9基编辑,绕过限制修改系统

Hung-Chun Lin¹, Wan-Chi Hsiao^2,3, Ya-Chen Hsu¹

¹Department of Chemistry, National Taiwan University, Taipei, Taiwan.

Applied and environmental microbiology

|March 10, 2025

概括

研究人员开发了一种CRISPR-Cas9细胞因子基编辑系统 (cBEST),用于在Bifidobacterium中高效的基因组编辑,这是一个关键的益生菌属. 绕过限制修改系统显著增强编辑,使新的治疗应用.

科学领域:

微生物学微生物学
分子生物学分子生物学
基因工程是一种基因工程.

背景情况:

双杆菌物种是重要的肠道微生物群,具有显著的益生菌和治疗潜力.
有限的遗传工具和普遍的限制修饰 (RM) 系统阻碍了对Bifidobacterium的遗传操纵.
有效的基因组编辑对于释放Bifidobacterium对人类健康的全部潜力至关重要.

研究的目的:

建立一个便携式的CRISPR-Cas9细胞酶基编辑系统 (cBEST),用于Bifidobacterium的基因组编辑.
研究RM系统对编辑效率的影响,并制定克服这些障碍的策略.
为了证明cBEST系统在多种Bifidobacterium物种中的可移植性和实用性.

主要方法:

开发和表征CRISPR-Cas9细胞因子基编辑器 (cBEST) 与各种促进器的等离子体.
在不同Bifidobacterium菌株和基因组环境中评估编辑效率.
破坏或绕过RM系统以增强基础编辑的策略.
在RM破坏的Bifidobacterium longum中应用cBEST,以简化基因组编辑.
测试cBEST在多种Bifidobacterium物种中对基因破坏的可移植性.

主要成果:

建立了一个便携式的CRISPR-Cas9细胞因子基编辑器 (cBEST) 系统,用于Bifidobacterium基因组编辑.
证明破坏或绕过RM系统可以显著提高编辑效率,特别是在具有挑战性的菌株和位置.
优化了cBEST等离子体设计,通过不同的促销元件来实现可调节的编辑效率.
简化了基因组编辑工作流程,使用RM破坏的Bifidobacterium longum菌株.
在使用相同的cBEST构造的多个Bifidobacterium物种中成功破坏了保存的代谢基因.

结论:

开发的cBEST系统为Bifidobacterium中基础编辑提供了一个高效且便携式的工具.
克服RM系统障碍对于推进Bifidobacterium的基因组编辑至关重要.
这种扩展的工具箱将加速对Bifidobacterium在宿主生理学和疾病中的作用的研究,为新的治疗应用铺平道路.

关键词:

这种细菌是Bifidobacterium.这就是CRISPR-Cas.通过DNA甲基化.代谢干扰代谢干扰限制修改系统的限制修改系统应变工程是一种应变工程.

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