Euphorbiaceae species are renowned not only for horticultural significance but for their production of numerous bicyclic diterpenes with antitumor and antiviral activities. However, the gene clusters responsible for the biosynthesis of these terpenes remain largely unidentified. We here initiated the construction of a comprehensive procedure for terpene gene clusters in Euphorbiaceae species. A total of 1824 candidate gene clusters with the range of 30–800 kb were identified across seven representative species including Ricinus communis, Hevea brasiliensis, Euphorbia peplus, Jatropha curcas, Manihot esculenta, Vernicia montana, and Vernicia fordii in Euphorbiaceae. The 16 high-confidence terpene gene clusters were ultimately pinpointed in Euphorbiaceae after satisfied the three stringent screening criteria: TPS/CYP pairwise relationship, copathway and coexpression patterns. Notably, the well-known casbene and casbene-derived diterpenoid gene cluster, involved in the biosynthesis of casbene, neocembrene, ingenanes, and jatrophanes, were identified. It was observed that casbene gene clusters were universally presented in Euphorbiaceae species, except M. esculenta. Among the casbene gene cluster, the alcohol dehydrogenase (ADH) was initially appeared, and neocembrene synthase is exclusively present in R. communis while absent in all the other species. These findings represent a significant step toward understanding the genetic basis of terpene biosynthesis in Euphorbiaceae species. Moreover, this knowledge on gene clusters responsible for the biosynthesis of pharmacologically relevant terpenes can serve as a theoretical foundation for future applications.

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