Introduction to Genome Editing Tool in Plants

Targeted genome editing has become a premier interest of researchers with the aim of gaining more insights into functional genomics and its applications for crop improvement. Conventional methods for genome editing based on sequence-specific nucleases such as zinc-finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs) are complex and difficult to engineer. A new genome editing platform based on bacterial Type II cluster regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated nuclease (Cas) has emerged as a powerful tool with high affordability, robustness, and simple engineering. CRISPR/Cas9 is a RNA-guided tool that introduces site-specific double-stranded breaks (DSBs) in the target genome using CRISPR-associated nuclease (Cas). DSBs are immediately restored through a host repair system, mainly non-homologous end joining (NHEJ) or homologous recombination (HR), which leads to insertions/deletions mutations (indel) in the defined target. CRISPR/Cas has become an important tool for creating diverse and heritable modifications at the targeted sites in plant genome. However, the high specificity required for a target sequence prohibited the wide range applicability of this technology, which led the researchers to find alternative nucleases such as Cas9 from other bacteria, as well as Cas12a and Cas13. Also, new tools are developed that rely on inactive or dysfunctional Cas proteins such as base editors and prime editors. With such technologies, genome editing has opened new paths for sustainable agriculture. This chapter provides a brief introduction to the mechanism and architecture of the CRISPR/Cas system, its progress, and its application in plants. In addition, the most recent application of Cas9 and its variants for the ectopic regulation of gene expression in plants are highlighted in this chapter.