Abstract
Betalains differ from anthocyanins, and they cannot coexist in the same plant under natural conditions. The L-DOPA 4,5-dioxygenase encoded by the DODA gene is a key step in the pathway of betalain biosynthesis in Caryophyllales plants. Amaranth is an important resource for the study and extraction of betalains. In order to clarify the function of AtrDODA family genes in betalain biosynthesis, we screened out three AtrDODA family gene members associated with betalains based on a genome database and RNA-seq databases of Amaranthus tricolor. Their characterization and expression pattern were further analyzed. The result of subcellular localization showed that all three AtrDODA members were located in the nucleus. Betacyanin and betaxanthin were promoted by paclobutrazol treatment in the leaves and stems of 'Suxian No.1' (red), while they were inhibited by gibberellin and darkness, which were consistent with the gene expression pattern of AtrDODAs. After heterologous transformation of the AtrDODA1-1 promoter into tobacco with GUS staining analysis, the promoter activity of AtrDODA1-1 of 'Suxian No.1' (red) amaranth was significantly higher than that of 'Suxian No.2' (green) amaranth. Furthermore, we analyzed the promoter activity of AtrDODA1-1 by GUS staining and qRT-PCR after sprayed exogenous MeJA and GA(3) on the AtrDODA1-1 promoter transformed tobacco plants. The results showed that AtrDODA1-1 responded to plant hormones. This study could lay a foundation for revealing the biological functions of the amaranth DODA gene family, and provide new clues for the molecular mechanism of betalain synthesis.