A haplotype-resolved chromosome-level genome of the hexaploid medicinal plant Plantago asiatica reveals subgenome asymmetry in bioactive compound biosynthesis

Plantago asiatica L. (Cheqian) is a traditional medicinal plant widely used in East Asia, whose seeds and whole plant serve as raw materials for multiple bioactive compounds, including geniposidic acid, acteoside, and plantamajoside. However, the lack of a high-quality reference genome has hindered molecular studies on the biosynthesis of medicinal compounds and polyploid evolution. Here, we report a haplotype-resolved, chromosome-level genome assembly of P. asiatica ‘Jiangcheqian’, a cultivated variety with superior agronomic and medicinal traits. The assembled genome spans 4.31 Gb with a scaffold N50 of 112.88 Mb, and 95.97% of the sequences are anchored to 36 pseudochromosomes, achieving a BUSCO completeness of 96.65%. The expansion of Copia retrotransposons is identified as a major contributor to the large genome size. Comparative genomic analyses reveal that the hexaploid genome comprises three distinct subgenomes (A, B and C), each represented by a pair of highly collinear haplotypes. Phylogenomic analysis combined with the distribution of synonymous substitution rates ( Ks ) indicates that the P. asiatica genome has experienced at least two whole-genome duplication (WGD) events; subgenome B and C underwent the most recent divergence event. Subgenome-specific functional divergence is suggested from the uneven distribution of unique gene families and from contrasting patterns of gene family expansion and contraction. The core gene families (UGT, 4CL, CYP450) involved in the biosynthesis of geniposidic acid, acteoside, and plantamajoside exhibit asymmetric gene retention across subgenomes and tissue-dependent subgenome-biased expression patterns, based on the analysis of gene abundance and transcript abundance. This high-quality chromosome-level genome provides a fundamental resource for deciphering polyploid evolution of the genus Plantago, unraveling the biosynthetic mechanisms of medicinal compounds, and accelerating molecular breeding of P. asiatica and allied medicinal Plantaginaceae taxa.