[Cloning and temporal-spatial expression analysis of dfr gene from Scutellaria baicalensis with different colors]

Dihydroflavanol-4-reductase (Dfr) is a key enzyme that regulates the synthesis of anthocyanin and proanthocyanidin in the flavonoid biosynthesis pathway. To investigate the difference of dfr gene in Scutellaria baicalensis Georgi with different colors in the same ecological environment, three complete full-length sequences of dfr gene were cloned from the cDNA of S. baicalensis with white, purple-red and purple colors using homologous cloning and RACE techniques. The three genes were named Sbdfr1, Sbdfr2 and Sbdfr3, respectively, and their corresponding structures were analyzed. The results showed that all three Dfr proteins have highly conserved NADPH binding sites and substrate-specific binding sites. Phylogenetic analysis showed that they are closely related to that of the known S. viscidula (ACV49882.1). Analysis of key structural domains and 3D models revealed differences in the catalytically active regions on the surface of all three Dfr proteins, and their unique structural characteristics may provide favorable conditions for studying the substrate specificity of different Dfr proteins. qRT-PCR analysis shows that dfr was expressed at different level in all tissues except the roots of S. baicalensis in full-bloom. During floral development, the expression level of dfr in white and purple-flowered Scutellaria showed an overall upward trend. In purple-red-flowered Scutellaria, the expression first slowly increased, followed by a decrease, and then rapidly increased to the maximum. This research provides a theoretical basis for further exploring the mechanism and function of Dfr substrate selectivity, and are of great scientific value for elucidating the molecular mechanism of floral color variation in S. baicalensis.