Characterisation of Early Biosynthetic Steps of Atractylon via an Integrative Strategy.

Atractylodes lancea, a vital medicinal plant, is renowned for its bioactive sesquiterpenoids, particularly atractylon. Despite its therapeutic importance, the genetic and biosynthetic pathways underlying atractylon production remain insufficiently understood. In this study, we present a comprehensive chromosome-scale genome assembly of A. lancea with a contig N50 of 263 Mb, advancing the existing genomic knowledge with unprecedented detail. Utilising an integrative omics approach, we coupled genome resequencing across 245 diverse plant accessions, metabolomics and transcriptomics comparisons among different A. lancea genetic backgrounds to profile the sesquiterpenoid divergency and construct a biosynthetic map of the sesquiterpenoids. Metabolite-based genome-wide association studies (mGWAS) reveal significant genetic variations related to atractylon diversity, offering terpene synthase (TPS) gene AlTPSa47 may be involved. Our work further identified and characterised key genes within the TPS family. Catalytic characterisation showed that five of them processed FPP into 12 different sesquiterpenoids. Metabolic profiling determined that transgenic roots overexpressing AlTPSa47 not only increased β-elemene and germacrene B, two of its enzymatic products, but also enhanced atractylon, notably confirming AlTPSa47 is essential in the initial steps of atractylon biosynthesis through rigorous transgenic validation. Collectively, our findings provide foundational resources for advancing breeding programs and enhancing the production of therapeutic compounds in A. lancea, marking a significant leap forward in medicinal plant biotechnology.