Biotechnology Letters ( IF 2.7 ) Pub Date : 2021-09-17 , DOI: 10.1007/s10529-021-03177-z Guohui Xiao 1 , Xiangdong Fu 1 , Juanjuan Zhang 1 , Shuyan Liu 1 , Zhaoqin Wang 1 , Taosheng Ye 1 , Guoliang Zhang 1
Objective
With the widespread application of CRISPR/Cas9 gene editing technology, new methods are needed to screen mutants quickly and effectively. Here, we aimed to develop a simple and cost-effective method to screen CRISPR/Cas9-induced mutants.
Result
We report a novel method to identify CRISPR/Cas9-induced mutants through a DNA-guided Argonaute nuclease derived from the archaeon Pyrococcus furiosus. We demonstrated that the Pyrococcus furiosus Argonaute (PfAgo)-based method could distinguish among biallelic mutants, monoallelic mutants and wild type (WT). Furthermore, this method was able to identify 1 bp indel mutations.
Conclusion
The PfAgo-based method is simple to implement and can be applied to screen biallelic mutants and mosaic mutants generated by CRISPR-Cas9 or other kinds of gene editing tools.
中文翻译:
通过 DNA 引导的 Argonaute 核酸酶快速且经济高效地筛选 CRISPR/Cas9 诱导的突变体
客观的
随着CRISPR/Cas9基因编辑技术的广泛应用,需要新的方法来快速有效地筛选突变体。在这里,我们旨在开发一种简单且具有成本效益的方法来筛选 CRISPR/Cas9 诱导的突变体。
结果
我们报告了一种新方法,通过源自古细菌Pyrococcus furiosus的 DNA 引导的 Argonaute 核酸酶来识别 CRISPR/Cas9 诱导的突变体。我们证明了基于Pyrococcus furiosus Argonaute (PfAgo) 的方法可以区分双等位基因突变体、单等位基因突变体和野生型 (WT)。此外,该方法能够识别 1 bp 插入缺失突变。
结论
基于 PfAgo 的方法实施简单,可用于筛选由 CRISPR-Cas9 或其他类型的基因编辑工具生成的双等位基因突变体和镶嵌突变体。