Prime-editing systems have the capability to perform efficient and precise genome editing in human cells. In this study, we first developed a plant prime editor 2 (pPE2) system and test its activity by generating a targeted mutation on an HPT^-ATG reporter in rice. Our results showed that the pPE2 system could induce programmable editing at different genome sites. In transgenic T₀ plants, pPE2-generated mutants occurred with 0%–31.3% frequency, suggesting that the efficiency of pPE2 varied greatly at different genomic sites and with prime-editing guide RNAs of diverse structures. To optimize editing efficiency, guide RNAs were introduced into the pPE2 system following the PE3 and PE3b strategy in human cells. However, at the genomic sites tested in this study, pPE3 systems generated only comparable or even lower editing frequencies. Furthemore, we developed a surrogate pPE2 system by incorporating the HPT^-ATG reporter to enrich the prime-edited cells. The nucleotide editing was easily detected in the resistant calli transformed with the surrogate pPE2 system, presumably due to the enhanced screening efficiency of edited cells. Taken together, our results indicate that plant prime-editing systems we developed could provide versatile and flexible editing in rice genome.

Prime-editing 系统能够在人类细胞中进行高效精确的基因组编辑。在这项研究中，我们首先开发了植物主要编辑器 2 (pPE2) 系统，并通过在水稻中的 HPT- ^ATG报告^基因上产生靶向突变来测试其活性。我们的结果表明 pPE2 系统可以在不同的基因组位点诱导可编程编辑。在转基因 T ₀在植物中，pPE2 产生的突变体以 0%–31.3% 的频率发生，这表明 pPE2 的效率在不同的基因组位点和不同结构的引物编辑引导 RNA 的差异很大。为了优化编辑效率，按照人类细胞中的 PE3 和 PE3b 策略，将向导 RNA 引入 pPE2 系统。然而，在本研究中测试的基因组位点，pPE3 系统仅产生相当甚至更低的编辑频率。此外，我们通过结合 HPT ^-ATG开发了一种替代 pPE2 系统记者来丰富主要编辑的细胞。在用替代 pPE2 系统转化的抗性愈伤组织中很容易检测到核苷酸编辑，这可能是由于编辑细胞的筛选效率提高了。总之，我们的结果表明，我们开发的植物主要编辑系统可以在水稻基因组中提供多功能和灵活的编辑。