The conversion of flavonoid aglycones to their glycosides by plant glycosyltransferases may affect a wide range of outcomes, including stability, solubility and bioavailability. Scutellaria barbata, rich in flavonoid glycosides, is widely used as a traditional Chinese herbal medicine. In this study, a flavonoid glycosyltransferase cDNA (SbUGT) and its promoter from S. barbata were cloned and characterized as a flavonoid glycosyltransferase using whole-cell biotransformation. Fragments of different lengths of the 5'-flanking region of the SbUGT gene were fused to the beta-glucuronidase (GUS) gene and analyzed with transgenic Arabidopsis plants using histochemical and fluorometric assays. GUS activity in transgenic plants carrying the SbP-850U construct (-850 to +86 relative to the transcription start site) displayed the highest level and was enhanced by salt and methyl jasmonate, similar to the expression patterns of the endogenous SbUGT. GUS activity disappeared when the promoter was deleted to -98, and deletion analyses indicated the existence of positive and negative regulatory element(s). Unexpectedly, plants carrying the construct SbP-102U (-102 to +86) exhibited strong GUS activity exclusively in the roots. Our experiments revealed that the specific expression is mediated by different promoter regions and the unique region driving root-preferred expression can be used as a root-specific promoter.