A haplotype-resolved lychee pangenome uncovers structural and haplotype variations associated with flowering time divergence and fruit sugar composition

Lychee is a representative tropical-subtropical fruit species of southern China and is widely appreciated for its unique and characteristic flavor. Pangenome provides a powerful framework for capturing genetic variation and elucidating the genomic basis of phenotypic diversity in economically important crops. Here, we polished the lychee reference genome by reassembling with advanced sequencing technologies and manually curating gene models with transcriptomic support. We confirmed the hybrid origin of the elite lychee cultivar 'Feizixiao', and found that recombination has contributed to its superior fruit quality and robust environmental adaptability. We also generated haplotype-resolved genomes for 12 representative lychee accessions and constructed the first lychee pangenome, revealing extensive structural variations (SVs) and widespread gene copy number variations across the species that affect a substantial proportion of genes. Leveraging the pangenome together with resequencing data from 314 lychee accessions, we uncovered genomic variants underlying key agronomic traits in lychee. A ∼10 kb structural variant located in the promoter region of LcFT3 that affects transcriptional regulation of the CO-NF complex, providing a mechanistic explanation for flowering time divergence within the population. In addition, SVs at the LcSAI locus, like SNPs, are significantly associated with fruit sugar content. We characterized both coding and cis-regulatory variants of LcSAI, revealing their combined effects on allelic enzyme activity and gene expression that together shape fruit sugar composition. Collectively, this study establishes a comprehensive genomic resource for lychee and demonstrates that structural and haplotype-level variations captured by a pangenome underlie key developmental and metabolic traits in perennial fruit crops.