Identification of CRISPR-Induced Mutations in Plants: with a Focus on the Next-Generation Sequencing Assay

As the CRISPR/Cas system is becoming an indispensable and most effective gene editing tool in eukaryotic cells, the need for efficient analysis of the gene editing outcomes also grows. Notably, ‘in planta’ gene editing, carried out by the Cas/sgRNA transgene after Agrobacterium-mediated plant transformation, demands several effective strategies to detect CRISPR gene-edited mutations. It is because it lacks “cell screening” steps which are commonly included in the CRISPR gene editing in most mammalian or plant protoplast systems. In this review, we summarized and compared several CRISPR-mutant screening assays to facilitate the identification of correctly edited alleles. We especially described the Next-Generation Sequencing (NGS)-based analysis in detail. We hope that many labs can employ this strategy as the NGS screening is precise and quantitative and can be applied to large-scale screening while still being quantitative and accurate.