Abstract
To elucidate flavonoid biosynthesis in gentian flowers, we attempted molecular cloning of R2R3-MYB genes expressed in flower petals by degenerate PCR technique. Sequence analysis showed that the obtained fragments were classified into 24 groups. Among them, four sequences were abundant and full length cDNA sequences were determined with the RACE technique. The isolated MYB cDNAs were designated as GtMYB2a, GtMYB2b, GtMYB3 and GtMYB4. Phylogenic analysis showed that GtMYB3 and GtMYB4 belong to a group regulating phenylpropanoid and the anthocyanin biosynthetic pathway. The deduced amino acid sequence of the GtMYB4 showed a high homology with maize P and Arabidopsis MYB12. A transient expression assay using gentian petals confirmed that GtMYB4 could activate both promoters of chalcone synthase (GtCHS) and flavone synthase (GtFNSII) genes. Northern blot analysis indicated that GtMYB4 is expressed in early developmental stages of the flower. GtMYB4-expressing transgenic tobacco plants showed increased flavonol and decreased anthocyanin accumulation in flowers.
