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克里斯普尔/Cas9系统是一个适合用于丝状真菌Monascus pilosus的基因向编辑工具

Yunxia Gong¹, Shengfa Li¹, Qianrui Liu¹

¹College of Food Science and Technology, Huazhong Agricultural University, Wuhan, 430070, China.

Applied microbiology and biotechnology

|January 19, 2024

概括

我们为Monascus pilosus开发了CRISPR/Cas9基因编辑系统 (CGES),实现了比传统方法更高的基因替换频率. 这一进步增强了Monascus pilosus基因功能探索和Monacolin K生产.

科学领域:

菌类学菌类学是指菌类学.
分子生物学分子生物学
生物技术是生物技术.

背景情况:

(Monascus pilosus) 是一种传统上用于生产降脂化合物莫纳科林K (MK) 的真菌.
探索M. pilosus的基因组可以揭示改善化合物生产的新基因.
有效的基因操纵工具对于加速M. pilosus.基因功能发现至关重要.

研究的目的:

开发一个高效的CRISPR/Cas9基因编辑系统 (CGES) 对Monascus pilosus.
为了比较CGES的基因替代频率 (GRFs) 与Agrobacterium tumefaciens介导的转化 (ATMT).
为了研究DNA损伤反应 (DDR) 基因 (mpclr4,mpdot1,mplig4) 在M. pilosus中的作用.

主要方法:

开发了一种准备适用于M. pilosus.中CGES的原生质的协议.
同时转化载体和捐赠者DNA成为M. pilosus原生菌.
对三个DDR基因 (mpclr4,mpdot1,mplig4) 的CGES和ATMT的GRF进行比较.

主要成果:

在M. pilosus.中,CGES的GRF大约是ATMT的五倍.
无活化mpclr4增强了DDR,并增加了对DNA损伤剂的耐受性.
无活化mpdot1和mplig4阻断了DDR通路并降低了DNA损伤耐受性.
与野生类型相比,Δmpclr4菌株的莫纳科林K (MK) 产量增加了52.6%.

结论:

与ATMT相比,CGES是M. pilosus的优越基因编辑工具.
在M. pilosus.中,DNA损伤反应基因起着至关重要的作用.
无活化mpclr4提高了MK的生产,为改善工业应用提供了一种战略.

关键词:

这是一种由Agrobacterium tumefaciens介导的转化.这是一个CRISPR/Cas9系统.对DNA损伤的反应反应基因替代的频率是基因替代的频率.产生MK的能力.一个叫做Monascus的物种.

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