Efficient hairy root induction system of Astragalus membranaceus and significant enhancement of astragalosides via overexpressing AmUGT15.

key messageAstragalus membranaceus hairy roots induced by direct injection of Rhizobium rhizogenes with AmUGT15 overexpressing genes into the stem explants demonstrate enhanced astragaloside biosynthesis Astragalus membranaceus is a widely used medicinal plant, which has important economic, ecological, medicinal, and ornamental values for accumulating various triterpene saponins named astragalosides in roots. Although the hairy root culture technique has been established in A. membranaceus, the molecular regulation of metabolic pathways for improving astragaloside contents was not reported. In this study, an efficient hairy root induction method was established in A.membranaceus by directly injecting Rhizobium rhizogenes into the stem, with an induction rate of up to 80.1%. We improved the production of astragaloside in hairy roots by overexpressing AmUGT15, a 3-O-glucosyltransferase catalyzed xylosylation at C3-OH. The fluorescence microscopy observation revealed that the AmUGT15 fused with DsRed report gene constructed in T-DNA region was overexpressed in hairy roots, and the maximum biomass of hairy roots was measured on the 28th day of cultivation. HPLC analysis confirmed the total amount of astragalosides produced by AmUGT15 overexpressing hairy roots is 4.2 times higher than the non-transgenic control group. Our study proposed an effective method for astragalosides production in A. membranaceus hairy roots via metabolic engineering.