Identification and Characterization of Flavonoid Biosynthetic Gene Families in <i>Paeonia</i> Species and Their Roles in Stamen Petalization of <i>Paeonia lactiflora</i>

Flavonoid biosynthesis is proposed to play a critical role in floral organ development in <i>Paeonia</i> species. However, its specific involvement in stamen petalization remains unclear. This study identified and characterized 13 gene families related to flavonoid biosynthesis across four <i>Paeonia</i> species. Comparative and phylogenetic analysis revealed that most flavonoid biosynthesis-related genes experience lineage-specific expansion in <i>P. ludlowii.</i> Genes belonging to the same family were commonly clustered on chromosomes and displayed highly conserved domain and motif compositions. The <i>cis-element</i> analysis identified <i>Cis</i>-acting elements associated with light, hormonal, and stress responses, implicating their regulatory roles in flavonoid biosynthesis. To further investigate the role of these genes in stamen petalization of <i>P. lactiflora</i>, expression profiling analyses were performed on ‘Fen Yu Nu’ (normal stamens) and ‘Lian Tai’ (petaloid stamens) cultivars using transcriptomic data released previously. Three quercetin-related genes revealed distinct stage-specific patterns in ‘Fen Yu Nu’ and ‘Lian Tai’. Notably, <i>PlaF3’H03</i> exhibited significant upregulation during petaloid stamen development in ‘Lian Tai’, suggesting its role in stamen transformation. Molecular docking identified <i>PlaF3’H07</i> as a key enzyme with strong substrate-binding affinity (ΔG = −4.7 kcal/mol), supporting its catalytic function in quercetin synthesis. The expression pattern of key flavonoid biosynthetic genes was also confirmed across three developmental stages of floral buds by real-time quantitative PCR. This study provides insights into the genetic basis underlying stamen petalization in <i>P. lactiflora</i> and offers potential targets for genetic improvement of floral traits in <i>Paeonia</i> and other ornamental plants.