Genome Engineering of Eubacterium limosum Using Expanded Genetic Tools and the CRISPR-Cas9 System.,ACS Synthetic Biology

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Genome Engineering of Eubacterium limosum Using Expanded Genetic Tools and the CRISPR-Cas9 System.
ACS Synthetic Biology ( IF 3.7 ) Pub Date : 2019-08-12 , DOI: 10.1021/acssynbio.9b00150
Jongoh Shin ₁ , Seulgi Kang ₁ , Yoseb Song ₁ , Sangrak Jin ₁ , Jin Soo Lee ₁ , Jung-Kul Lee ₂ , Dong Rip Kim ₃ , Sun Chang Kim _{1,

4} , Suhyung Cho ₁ , Byung-Kwan Cho _{1,

4}

Affiliation

Eubacterium limosum is one of the important bacteria in C1 feedstock utilization as well as in human gut microbiota. Although E. limosum has recently garnered much attention and investigation on a genome-wide scale, a bottleneck for systematic engineering in E. limosum is the lack of available genetic tools and an efficient genome editing platform. To overcome this limitation, we here report expanded genetic tools and the CRISPR-Cas9 system. We have developed an inducible promoter system that enables implementation of the CRISPR-Cas9 system to precisely manipulate target genes of the Wood-Ljungdahl pathway with 100% efficiency. Furthermore, we exploited the effectiveness of CRISPR interference to reduce the expression of target genes, exhibiting substantial repression of several genes in the Wood-Ljungdahl pathway and fructose-PTS system. These expanded genetic tools and CRISPR-Cas9 system comprise powerful and widely applicable genetic tools to accelerate functional genomic study and genome engineering in E. limosum.

中文翻译：

使用扩展遗传工具和 CRISPR-Cas9 系统对真杆菌进行基因组工程。

利莫苏真杆菌是 C1 原料利用以及人类肠道微生物群中的重要细菌之一。尽管近年来，柠檬桉在全基因组范围内引起了广泛关注和研究，但柠檬桉系统工程的瓶颈是缺乏可用的遗传工具和高效的基因组编辑平台。为了克服这一限制，我们在这里报告了扩展的遗传工具和 CRISPR-Cas9 系统。我们开发了一种诱导型启动子系统，能够实现 CRISPR-Cas9 系统以 100% 的效率精确操纵 Wood-Ljungdahl 途径的靶基因。此外，我们利用 CRISPR 干扰的有效性来减少靶基因的表达，显示出对 Wood-Ljungdahl 途径和果糖-PTS 系统中多个基因的显着抑制。这些扩展的遗传工具和 CRISPR-Cas9 系统包含强大且广泛适用的遗传工具，可加速柠檬黄的功能基因组研究和基因组工程。

更新日期：2019-08-13

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