基于CRISPR/Cas9的基因组编辑用于Kluyveromyces lactis中的蛋白质表达和分泌
Lingtong Liao1,2, Xiuru Shen1,2, Zhiyu Shen1,2
1Science Center for Future Foods, Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu 214122, China.
ACS synthetic biology
|June 13, 2024
概括
我们为Kluyveromyces lactis开发了CRISPR/Cas9基因组编辑,增强了其作为生物制造平台的潜力. 这种工具可以有效地编辑基因,以改善酵母细胞工厂中蛋白质和化学品的生产.
科学领域:
- 工业生物技术 工业生物技术
- 微生物工程 微生物工程
- 合成生物学 合成生物学
背景情况:
- 乳酪酵母是一种有前途的酵母,可用于工业生物技术.
- 有限的遗传工具阻碍了其作为生物制造平台的发展.
研究的目的:
- 在K. lactis.中开发和应用CRISPR/Cas9基因组编辑.
- 建立单个和多个基因编辑的高效方法.
- 利用基因组编辑来增强蛋白质的生产和特征分泌途径.
主要方法:
- 在K. lactis.中实施CRISPR/Cas9系统.
- 破坏KU80基因以提高编辑效率.
- 为同时进行多基因编辑,优化sgRNA传递.
- 编辑事件的彩色指示器辅助选.
- 多站点集成和基因淘汰策略.
主要成果:
- 通过破坏KU80.0,实现了80%的单基因编辑效率.
- 在两基因和三基因编辑方面分别达到73.3%和36%的效率.
- 通过多站点集成成功增强乳糖酶的产生.
- 通过组合基因淘汰来表征乳糖酶细胞外分泌.
结论:
- CRISPR/Cas9是K. lactis基因组工程的一个强大的工具.
- 开发的方法使得高效的单个和多重基因编辑成为可能.
- 这项技术使K. lactis成为生物生产的细胞工厂.
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