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多复合的CRISPR-Cas9协议用于大型转基因集成到Schistosoma mansoni基因组中

Wannaporn Ittiprasert¹, Max M Moescheid², Victoria H Mann¹

¹Department of Microbiology, Immunology and Tropical Medicine, School of Medicine and Health Sciences, George Washington University, Washington, DC 20037, USA.

|February 14, 2024

概括

这项研究引入了一种多重化的CRISPR-Cas9方法,用于将大型DNA序列集成到Schistosoma mansoni基因组中. 这一进步使得在研究和治疗开发中能够有效地对瘤进行基因工程.

科学领域:

遗传学和基因组学在
分子生物学分子生物学
寄生虫学的寄生虫学

背景情况:

克里斯普尔-Cas9技术允许精确的基因组编辑.
以前的研究表明,使用NHEJ和HDR途径在Schistosoma mansoni中显示了CRISPR-Cas9.
大规模的转基因整合是S. mansoni的一个重大挑战.

研究的目的:

开发和展示一个多重化的CRISPR-Cas9协议,用于将大型转基因整合到Schistosoma mansoni基因组中.
优化将CRISPR组件和捐赠者DNA输送到分裂体卵中的方法.
为了促进S. mansoni的先进基因操纵.

主要方法:

制备多重核核蛋白复合物 (RNP).
设计和制备具有5'酸修饰的双链供体DNA.
通过正方形波电穿孔将RNP和捐赠者DNA引入分裂体卵.

主要成果:

成功展示了用于大型转基因集成的多重CRISPR-Cas9协议.
关于RNP和供体DNA的准备和输送的详细程序.
在S. mansoni.中建立了一种高效基因工程的方法.

结论:

提出的协议使得高效的,大规模的转基因集成可以在Schistosoma mansoni.
这种方法提升了S. mansoni研究的基因操纵能力.
该协议为未来关于瘤生物学和控制的研究提供了基础.

关键词:

克里斯普尔是什么意思?克里斯普尔是什么意思?基因表达基因表达模型生物模型生物分子生物学分子生物学

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CRISPR/Cas9 Genome Editing

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The CRISPR-Cas system serves as a bacterial defense mechanism against invading genetic elements such as viruses and plasmids, forming the foundation for its adaptation as a powerful genome-editing tool. Originally discovered in prokaryotes, this system has been repurposed to revolutionize genetic engineering across a wide range of organisms, including plants, animals, and humans. The core component, Cas9, is an endonuclease derived from Streptococcus pyogenes, capable of introducing...