Expanding the plant genome editing toolbox with recently developed CRISPR-Cas systems

Abstract

Since its first appearance, CRISPR-Cas9 has been developed extensively as a programmable genome-editing tool, opening a new era in plant genome engineering. However, CRISPR-Cas9 still has some drawbacks, such as limitations of the protospacer-adjacent motif (PAM) sequence, target specificity, and the large size of the cas9 gene. To combat invading bacterial phages and plasmid DNAs, bacteria and archaea have diverse and unexplored CRISPR-Cas systems, which have the potential to be developed as a useful genome editing tools. Recently, discovery and characterization of additional CRISPR-Cas systems have been reported. Among them, several CRISPR-Cas systems have been applied successfully to plant and human genome editing. For example, several groups have achieved genome editing using CRISPR-Cas type I-D and type I-E systems, which had never been applied for genome editing previously. In addition to higher specificity and recognition of different PAM sequences, recently developed CRISPR-Cas systems often provide unique characteristics that differ from well-known Cas proteins such as Cas9 and Cas12a. For example, type I CRISPR-Cas10 induces small indels and bi-directional long-range deletions ranging up to 7.2 kb in tomatoes (Solanum lycopersicum L.). Type IV CRISPR-Cas13 targets RNA, not double-strand DNA, enabling highly specific knockdown of target genes. In this article, we review the development of CRISPR-Cas systems, focusing especially on their application to plant genome engineering. Recent CRISPR-Cas tools are helping expand our plant genome engineering toolbox.