The genome analysis of Tripterygium wilfordii reveals TwCYP712K1 and TwCYP712K2 responsible for oxidation of friedelin in celastrol biosynthesis pathway

is a Traditional Chinese Medicine (TCM) from family Celastraceae and celastrol is one of the strongest active ingredients belonging to friedelane-type pentacyclic triterpenoid, which has a large clinical application value of anti-tumor, immunosuppression, and obesity treatment. The first committed biosynthesis step of celastrol is the cyclization of 2, 3-oxidosqualene to friedelin, catalyzed by the oxidosqualene cyclase, while the rest of this pathway is still unclear. In this study, we reported a reference genome assembly of with high-quality annotation by using a hybrid sequencing strategy (Nanopore, Bionano, Illumina HiSeq, and Pacbio), which obtained a 340.12 Mb total size and contig N50 reached 3.09 Mb. In addition, we successfully anchored 91.02% sequences into 23 pseudochromosomes using Hi-C technology and the super-scaffold N50 reached 13.03 Mb. Based on integration genome, transcriptom and metabolite analyses, as well as and enzyme assays, two CYP450 genes, and have been proven for C-29 position oxidation of friedelin to produce polpunonic acid, which clarifies the second biosynthesis step of celastrol. Syntenic analysis revealed that 1 and derived from the common ancestor. These results have provided insight into illustrating pathways for both celastrol and other bioactive compounds found in this plant.