High level production of δ-guaiene, a bicyclic sesquiterpene accumulated in agarwood, by co-expression of δ-guaiene synthase and farnesyl diphosphate synthase genes in tobacco BY-2 cells

Two genes encoding δ-guaiene synthase (GS) and farnesyl diphosphate synthase (FPS) involved in δ-guaiene biosynthesis in Aquilaria microcarpa were co-expressed in tobacco (Nicotiana tabacum) BY-2 cells by Agrobacterium-mediated transformation. GC-MS analysis revealed that the transformed tobacco cells liberated δ-guaiene, and the compound was found in the headspace of the culture but not accumulated either in the medium or in the cells. Tobacco cells transformed by solely GS produced 0.2 mg δ-guaiene /L culture, however, concentration of the compound elevated to 0.6 – 5.9 mg/L when GS and FPS were co-expressed in the cells. The stirring efficiency of the cell suspension was improved by the reduction of the culture volume in the vials, and this resulted in an appreciable increase in δ-guaiene content to the level of 102 mg/L culture. Addition of mevalonolactone as the precursor markedly activated δ-guaiene production, and content of the compound elevated to more than 400 mg/L culture. These results strongly suggested that tobacco BY-2 is a suitable host to construct the bio-production system of sesquiterpene compounds, and co-expression of FPS and appropriate sesquiterpene synthase genes in the cells should be the good strategy to establish the highly productive platform of this class of compounds.