Vine tea (Nekemias grossedentata) is a dual-purpose medicinal and edible liana with a documented history of consumption in China spanning millennia. It has been extensively utilized among ethnic minority groups, including the Tujia, Yao, and Dong communities, for at least 700–1000 years, where it is traditionally revered as the ‘Immortal Herb’ or ‘Longevity Tea’. This study reports the haplotype-resolved chromosome-scale genomes of two major cultivated diploid vine tea accessions (N. grossedentata, 2n = 40). Phylogenetic analysis revealed that N. grossedentata diverged from Cissus rotundifolia ~26.27 million years ago (MYA) and from Vitis vinifera around 17.30 MYA. Comparative genomic analysis within the genus uncovered species-specific evolutionary patterns. Furthermore, we constructed a pan-genome encompassing 39 vine tea cultivars and characterized structural variations among cultivated varieties. Correlation analysis between dihydromyricetin (DMY) content and leaf transcriptomes across these cultivars identified ~1 kb presence/absence variations (PAVs) associated with the expression of F3′5′H, a gene critical for DMY biosynthesis in vine tea. Collectively, this genomic resource provides a valuable foundation for advancing herbal crop breeding and development, while offering insights into the biosynthetic pathways underlying specialized metabolism in Vitaceae.

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