Abstract
Peach (Prunus persica) is an economically important fruit crop globally and an excellent material for genomic studies. While considerable progress has been made in unveiling trait-associated genes within cultivars and wild relatives, certain novel genes controlling valuable traits in peach landraces, such as the red-flowering gene, remained unclear. In this study, we sequenced and assembled the diploid genome of the red-flower landrace 'Yingzui' (abbreviated as 'RedY'). Multi-omics profiling of red petals of 'RedY' revealed the intensified red coloration associated with anthocyanins accumulation and concurrent decline in flavonols. This phenomenon is likely attributed to a natural variant of Flavonol Synthase (FLS) harboring a 9-bp exonic insertion. Intriguingly, the homozygous allelic configurations of this FLS variant were only observed in red-flowered peaches. Furthermore, the 9-bp sequence variation tightly associated with pink/red petal color in genome-wide association studies (GWAS) of collected peach germplasm resources. Functional analyses of the FLS variant, purified from procaryotic expression system, demonstrated its diminished enzymatic activity in flavonols biosynthesis, impeccably aligning with the cardinal trait of red flowers. Therefore, the natural FLS variant was proposed as the best candidate gene for red-flowering trait in peach. The pioneering unveiling of the red-flowered peach genome, coupled with the identification of the candidate gene, expanded the knowledge boundaries of the genetic basis of peach traits and provided valuable insights for future peach breeding efforts.