Precise Editing of the Wx Gene Promoter via Prime Editing to Develop Novel Rice Germplasm with Low Amylose Content

Rice (Oryza sativa L.) is a staple food, and its eating quality is largely determined by amylose content (AC), which is controlled by the Wx gene. Current low-AC alleles often lead to excessive reductions, limiting their utility. The promoter region, particularly the pyrimidine-rich region (PRR) from -45 to -7, is hypothesized to fine-tune Wx expression, yet its functional motifs remain unexplored. In this study, we applied prime editing (PE) to precisely edit the Wx promoter in the japonica cultivar Huaidao 5. We generated two novel alleles, Wx^1T/32d (deleting PRR1 [-45 to -19] and PRR2 [-11 to -7]) and Wx^5d (deleting only PRR2), which are absent in natural germplasm. These edits resulted in significant, fine-tuned reductions in AC (3.95% and 2.77%, respectively) without affecting grain transparency. Our results identify PRR1 and PRR2 as important functional motifs whose deletion synergistically downregulates AC. Furthermore, the edited lines exhibited improved taste without compromising agronomic performance. This study not only provides novel insights into the regulatory motifs of the Wx promoter but also demonstrates the precision and potential of PE for improving rice grain quality.