Objective Evaluation of Nepeta cataria as a host with specific endogenous metabolite background for transient expression and metabolic engineering of secondary biosynthetic sequences. Results The reporter gene GFP::lic BM3 as well as three biosynthetic genes leading to the formation of the cannabinoid precursor olivetolic acid were adopted to the modular cloning standard GoldenBraid, transiently expressed in two chemotypes of N. cataria and compared to Nicotiana benthamiana . To estimate the expression efficiency in both hosts, quantification of the reporter activity was carried out with a sensitive and specific lichenase assay. While N. benthamiana exhibited lichenase activity of 676 ± 94 μmol g -1 s -1 (Gerasimenko et al. 2019), N. cataria cultivar ‘1000’, and the cultivar ‘Citriodora’ showed an activity of 37 ± 8 μmol g -1 s -1 and 18 ± 4 μmol g -1 s -1 , respectively. Further, combinatorial expression of genes involved in cannabinoid biosynthetic pathway acylactivating enzyme 1 ( aae1 ), olivetol synthase ( ols ) and olivetolic acid cyclase ( oac ) in N. cataria cv. resulted presumably in the in vivo production of olivetolic acid glycosides. Conclusion Nepeta cataria is amenable to Agrobacterium -mediated transient expression and could serve as a novel chassis for the engineering of secondary metabolic pathways and transient evaluation of heterologous genes.
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Geissler et al. (2023) studied this question.