Introduction of mutations in plants with prime editing.

Abstract

The emergence of a clustered regularly interspaced short palindromic repeats/CRISPR-associated proteins (CRISPR/Cas) system has had a revolutionary impact on plant biology. However, this system and further developed base editing are limited by their inherent imperfection. Prime editing, a just arrival technology based on CRISPR/Cas, can directly and precisely edit a specified DNA site without double strand breaks and donor DNA by integrating an engineered reverse transcriptase (RT) with a catalytically impaired Cas9 endonuclease and introducing genetic information into prime editing guide RNA (pegRNA). In addition, it has a wider range of editing types than base editing and can install all types of editing theoretically. Prime editing was originally developed in mammalian cells and has recently been applied to plants. Here, we describe the origin of prime editing and compare it with traditional CRISPR/Cas9 and base editing; then, we exemplify it in plants, including strategies and methods. Accordingly, we generate the overall procedures of prime editing to provide instructions for its application. Furthermore, we summarize its improvements in the approach, such as optimizing the length of a primer binding site and RT template, as well as pursuing an optimal nicking site in the unedited sequence. Finally, we discuss the potential impact on domestication and improvement of agricultural crops, sustainable utilization of medicinal plants, cultivation of varieties of horticultural plants, and revelation of the genetic code, in order to offer a reference for the further study and development of prime editing.