Identification and functional analysis of terpene synthases revealing the aroma variations among different...

In both natural and cultivated populations of Chrysanthemum indicum var. aromaticum (CIA), diploid and tetraploid germplasms coexist, yet the comparative aroma profiles and their biosynthetic mechanisms remain poorly characterized. This study used a combination of ultra-fast gas-phase electronic nose, widely targeted flavor-omics and sensory evaluation assays, and identified 15 core aroma compounds in diploid CIA and 10 in tetraploid CIA. Notably, the aroma differences between these two groups were primarily attributed to variations in terpenoids. Using comparative transcriptome analysis, 30 different terpene synthase (TPS) genes in floral tissues and 27 in leaf tissues across ploidy levels. Seven TPS genes were cloned and classified into two TPS subfamilies. Structure models of these seven TPSs were predicted, and molecular docking results indicated strong binding affinities with farnesyl diphosphate and geranyl diphosphate. In vitro enzymatic experiments demonstrated that six CiaTPS-a enzymes exhibited multifunctional catalytic activities, biosynthesizing floral and woody terpenoids including α-Terpineol, β-Bisabolene and (E)-β-Farnesene. In contrast, CiaTPS-b19 exhibited substrate specificity, exclusively converting geranyl diphosphate into (E)-β-Ocimene, β-Myrcene, and β-Ocimene. These findings provide crucial insights into the breeding and industrial applications of diploid and tetraploid CIA resources.