Rapid and cost-effective screening of CRISPR/Cas9-induced mutants by DNA-guided Argonaute nuclease

Xiao, Guohui; Fu, Xiangdong; Zhang, Juanjuan; Liu, Shuyan; Wang, Zhaoqin; Ye, Taosheng; Zhang, Guoliang

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. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s10529-021-03177-z.