Chromosome-level genome of Mosla chinensis from alpine ecotype provides insights into terpenoid biosynthesis and germplasm exploration.

Mosla chinensis is a high-value medicinal plant with scarce genomic resources for precision breeding. Here, we present the first chromosome-scale genome of an alpine ecotype (Baiyunjian Reserve, 1,611 m above sea level) assembled via SMRT sequencing and Hi-C technologies. 93.46% of sequences were anchored to 9 pseudochromosomes with 99.99% accuracy, resulting in the assembly of the 444.26 Mb genome (contig N50 = 41.38 Mb; 98.7% BUSCO). High proportion of repetitive sequences (56.90%) and species-specific genome duplication revealed the species-specific adaptations of this ecotype. Multi-tissue metabolomics identified flowers and leaves as biosynthetic hotspots for terpenoids especially γ-terpinene, thymol and carvacrol. Integrated with transcriptomics pinpointed MchTPS4 as the plastid-localized γ-terpinene synthase, while phylogenetic mining identified cytochrome P450 MchCYPs (MCH03g18750) and dehydrogenases MchSDRs (MCH04g17930/14560) as potential downstream enzymes. Functional validation confirmed that MchTPS4 was a monoterpene synthase while MchTPS2, MchTPS7, MchTPS9-11 were sesquiterpene synthases. Furthermore, metabolic engineering boosted the yield of γ-terpinene 3.45-fold to 5.09 mg/L. Biosynthetic gene clusters (BGCs) analyses uncovered 61 clusters with 11 associated with terpenoid metabolism. MchTPS2 co-localized in cluster 6 with two MchCYPs (MCH01g19200; MCH01g19220). Syntenic analysis revealed that MchTPS2 evolved as a single-copy, whereas MCH01g19200 underwent tandem duplication from M. chinensis to Salvia divinorum and S. hispanica. Phylogenomic analysis resolved M. chinensis as the sister species of Perilla frutescens (100% BS), diverging ~12.7 Mya in the mid-Miocene. Comparative genomics revealed a species-specific whole-genome duplication event post-dating divergence from Scutellaria baicalensis, driving syntenic retention of specialized metabolic genes. This study decodes the genetic basis of terpenoid biosynthesis in an extremophyte, providing resources for molecular breeding of traditional Chinese medical plant "Xiangru" and insights into Lamiaceae metabolic evolution.