多个sgRNAs的应用提高了在Arabidopsis中CRISPR/Cas9介导基因向的效率
Jing Li1,2, Dali Kong1,2, Yongping Ke1,2
1Shanghai Center for Plant Stress Biology, CAS Center for Excellence in Molecular Plant Sciences, Chinese Academy of Sciences, Shanghai, 200032, China.
BMC biology
|January 17, 2024
概括
通过放大 sgRNA 副本来增加 CRISPR/Cas9 双链断裂 (DSB),可以提高植物的基因向 (GT) 效率. 然而,使用多个sgRNA并不总是改善GT结果,有些人帮助其他人.
科学领域:
- 植物分子生物学 植物分子生物学
- 基因组工程是基因组工程.
- 生物技术是生物技术.
背景情况:
- 精确的基因向 (GT) 对于遗传基因组工程至关重要.
- 以前的研究将Cas9双链断裂 (DSB) 频率与植物中的GT效率联系起来.
- 同一个位置的DSB对GT的影响以及sgRNA复制数或多个sgRNA对GT效率的影响仍然未被探索.
研究的目的:
- 研究植物内源性位点的DSB和GT效率之间的关系.
- 为了确定增加sgRNA复制数或使用多个sgRNA是否会提高CRISPR/Cas9介导的GT效率.
- 阐明DSB频率在植物基因向中的作用.
主要方法:
- 在Arabidopsis thaliana和大米中针对CRISPR/Cas9介导的基因.
- 顺序转型战略. 顺序转型战略.
- 应用单个和多个sgRNAs具有不同的副本数.
- 在内源EMB2410和ROS1位点上分析DSB频率和GT效率.
主要成果:
- 在内生EMB2410和ROS1位点实现了精确的GT.
- 增加的sgRNA复制数增加了DSB和GT效率.
- 应用多个sgRNAs并没有持续提高GT效率.
- 观察到低效的sgRNAs可以促进其他sgRNAs的DSB活性.
结论:
- DSB效率是植物GT效率的关键决定因素,比突变模式更重要.
- 这项研究为sgRNAs,DSBs和GT之间的相互作用提供了新的见解.
- 这些发现有助于了解植物中CRISPR/Cas9介导GT的分子机制.
相关概念视频
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