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Genome Editing in Mammalian Cell Lines using CRISPR-Cas

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Published on: April 11, 2019

在Methanosarcina acetivorans中进行基因组编辑的CRISPR/Cas12a工具箱

Ping Zhu¹, Tejas Somvanshi¹, Jichen Bao¹

¹Laboratory of Biochemistry, Department of Bioproducts and Biosystems, School of Chemical Engineering, Aalto University, Espoo, Finland.

Frontiers in microbiology

|December 27, 2023

概括

研究人员开发了一种新的CRISPR-Cas12a工具箱,用于有效编辑甲原体中的基因组. 这种工具使得精确的基因删除和插入成为可能,进步了这些重要微生物的基因工程.

科学领域:

微生物学微生物学
合成生物学合成生物学
生物技术是生物技术.

背景情况:

甲原有古生物对全球碳循环至关重要,并具有生物燃料生产的潜力.
甲沙辛酸是一种成熟的模型生物体,用于甲原研究,因为它具有遗传可处理性.
之前在M. acetivorans中进行的基因组编辑依赖于CRISPR/Cas9系统.

研究的目的:

开发和描述一种易于使用的CRISPR-Cas12a工具箱,用于在Methanosarcina acetivorans中进行基因组编辑.
为了证明CRISPR-Cas12a在大型DNA删除和异质基因插入方面的效率.
为了展示CRISPR-Cas12a系统的多重基因组编辑能力.

主要方法:

开发一个CRISPR-Cas12a工具箱,针对富T的PAM序列 (5'-TTTV).
应用工具箱用于大规模的基因组删除 (高达3,500个基因组) 和基因插入.
利用Cas12a进行多重基因组编辑,同时针对多个染色体位点.

主要成果:

在CRISPR-Cas12a工具箱成功地促进了删除高达3,500 bp,包括整个frhADGB操作.
实现了异质基因插入,成功率超过80%.
多重编辑证明了高效率,双重删除 (100 bp) 和同时删除/插入 (100 bp删除,2,400 bp插入) 产生了高精度.

结论:

开发的CRISPR-Cas12a工具箱为Methanosarcina物种的基因组编辑提供了可靠和高效的方法.
该系统补充了现有的CRISPR/Cas9工具,扩大了基因工程对甲基生物的工具包.
增强的编辑功能促进了使用甲原体的生物技术应用.

关键词:

这就是CRISPR/Cas12a.甲沙辛酸乙基甲沙辛酸乙基甲沙辛酸基因组编辑基因组编辑甲氧化物是一种甲基化物.合成生物学合成生物学

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