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通过整合工程 Cas9 蛋白质来改善和简化 Yarrowia lipolytica 中的基因编辑

Baixi Zhang^1,2, Jiacan Cao¹

¹School of Food Science and Technology, Jiangnan University, Wuxi 214122, China.

Journal of fungi (Basel, Switzerland)

|January 22, 2024

概括

我们使用eSpCas9在Yarrowia lipolytica中改进了CRISPR基因编辑,显著提高了无需超生长步骤的效率. 这简化了这种有价值的油性酵母的代谢工程.

科学领域:

合成生物学合成生物学
微生物生物技术微生物生物技术

背景情况:

雅罗维亚 (Yarrowia lipolytica) 是生物炼油厂的关键油性酵母,在石油生产和蛋白质分泌方面表现出色.
对于Y. lipolytica而言,现有的CRISPR/Cas9系统需要超生长阶段,这限制了效率和实用性.
对Y. lipolytica的代谢工程需要高效和精简的基因组编辑工具.

研究的目的:

为了提高基因编辑效率和忠实在Yarrowia lipolytica.
在Y. lipolytica.中引入和优化eSpCas9基因的表达.
为了消除在Y. lipolytica基因组编辑中需要转化后的生长步骤.

主要方法:

将ESpCas9蛋白集成到Y. lipolytica基因组中.
通过使用不同的促进剂和生长周期,探索了最佳的eSpCas9基因表达.
在TRP1和LIP2位置评估基因编辑效率.
分析了酵母生长曲线和稀释试验.

主要成果:

实现显著提高基因编辑效率:TRP1的86.09% (从16.61%) 和LIP2的95.19% (从33.61%).
消除了为高编辑效率而需要耗时的增长步骤的要求.
观察到轻微的增长抑制与一致的eSpCas9表达,表明可诱导的促进剂用于未来的研究.

结论:

集成的eSpCas9系统简化了Y. lipolytica的基因组编辑,提高了效率.
这一进步增强了Y. lipolytica作为天然产品合成的底盘的潜力.
这些发现为改善其他微生物的基因组编辑提供了宝贵的见解.

关键词:

这就是CRISPR/Cas9的作用.雅罗维亚 (Yarrowia lipolytica) 是一个高脂的植物.基因组编辑基因组编辑

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