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基于CRISPR/Cas9的代多副本集成,以提高Saccharomyces cerevisiae中的代谢产量

Ximei Chen¹, Chenyang Li², Xin Qiu²

¹School of Biological Engineering, Dalian Polytechnic University, Dalian, 116034, China.

Synthetic and systems biotechnology

|March 28, 2025

概括

我们开发了Iterative Multi-copy Integration by Gene Editing (IMIGE) 系统,用于在酵母中高效地整合基因. 这种基于CRISPR/Cas9的方法显著提高了像ergothioneine和cordycepin这样有价值的化合物的产生.

科学领域:

合成生物学合成生物学
代谢工程是代谢工程.
微生物生物技术微生物生物技术

背景情况:

在Saccharomyces cerevisiae中高复制基因的整合对于高效的天然产品生物合成至关重要.
传统的多拷贝基因整合方法往往是低效和劳动密集的.

研究的目的:

开发一种新的,高效的系统,用于在S. cerevisiae中同时进行多拷贝基因集成.
简化菌株工程流程,以提高微生物生产.

主要方法:

使用CRISPR/Cas9技术开发基因编辑 (IMIGE) 系统的代多拷贝集成.
利用 δ 和 rDNA 重复序列进行同时集成.
在体内使用分裂标记器策略和Cas9-sgRNA表达载体进行捐赠者DNA组装.
基于与生长相关的表型的快速,代性选.

主要成果:

在两个查周期内,IMIGE系统实现了对欧戈氨酸 (105.31 mg/L) 和科迪塞宾 (62.01 mg/L) 的显著产量改善.
欧戈氨酸的产量增加了407.39%,而科尔迪塞宾的产量增加了222.13%,相比于发作性表达菌株.
证明了高通量应变工程的效率和可扩展性.

结论:

IMIGE系统为微生物菌株工程提供了一个高效和可扩展的平台.
这种方法简化了基因整合,减少了查需求,并提高了生物产品标位.
促进微生物生产有价值的自然产品和其他生物产品.

关键词:

这就是CRISPR/Cas9的作用.科尔迪斯的头脑.埃尔戈西奥尼因 (Ergothioneine) 是一种具有活力的物质.多复制集成的多复制集成这种植物是Saccharomyces cerevisiae.

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