Division of Applied Life Science (BK21 Four Program), Plant Molecular Biology and Biotechnology Research Center, Gyeongsang National University, Jinju, Korea.
Division of Life Science, Gyeongsang National University, Jinju, Korea.
Plant Biotechnol J. 2024 Jan;22(1):19-36. doi: 10.1111/pbi.14188. Epub 2023 Oct 4.
Prime editing (PE) technology utilizes an extended prime editing guide RNA (pegRNA) to direct a fusion peptide consisting of nCas9 (H840) and reverse transcriptase (RT) to a specific location in the genome. This enables the installation of base changes at the targeted site using the extended portion of the pegRNA through RT activity. The resulting product of the RT reaction forms a 3' flap, which can be incorporated into the genomic site through a series of biochemical steps involving DNA repair and synthesis pathways. PE has demonstrated its effectiveness in achieving almost all forms of precise gene editing, such as base conversions (all types), DNA sequence insertions and deletions, chromosomal translocation and inversion and long DNA sequence insertion at safe harbour sites within the genome. In plant science, PE could serve as a groundbreaking tool for precise gene editing, allowing the creation of desired alleles to improve crop varieties. Nevertheless, its application has encountered limitations due to efficiency constraints, particularly in dicotyledonous plants. In this review, we discuss the step-by-step mechanism of PE, shedding light on the critical aspects of each step while suggesting possible solutions to enhance its efficiency. Additionally, we present an overview of recent advancements and future perspectives in PE research specifically focused on plants, examining the key technical considerations of its applications.
碱基编辑(PE)技术利用扩展的碱基编辑向导 RNA(pegRNA)将包含 nCas9(H840)和逆转录酶(RT)的融合肽引导到基因组中的特定位置。这使得能够通过 RT 活性利用 pegRNA 的扩展部分在靶向位点安装碱基变化。RT 反应的产物形成 3' 突出端,该突出端可以通过一系列涉及 DNA 修复和合成途径的生化步骤整合到基因组位点中。PE 已证明其在实现几乎所有形式的精确基因编辑方面的有效性,例如碱基转换(所有类型)、DNA 序列插入和缺失、染色体易位和倒位以及基因组中安全港位点的长 DNA 序列插入。在植物科学中,PE 可以作为精确基因编辑的突破性工具,允许创建所需的等位基因来改良作物品种。然而,由于效率限制,其应用在双子叶植物中遇到了限制。在这篇综述中,我们讨论了 PE 的逐步机制,强调了每个步骤的关键方面,并提出了可能的解决方案来提高其效率。此外,我们还介绍了植物中 PE 研究的最新进展和未来展望,重点关注其应用的关键技术考虑因素。