| In recent years, the UFGT gene function of anthocyanins in color-leafed plants as corn, apple, grape, mulberry, tomato, carrot, cherry, strawberry, navel orange, and fruit tree leaves, has been investigated, but a little is known in the purple wheat seed coat. This paper verified the biological function of UFGT gene, and analyzed the expression pattern of UFGT gene in purple wheat seed coat. The study provides some data on anthocyanin biosynthesis in the purple wheat seed coat.UFGT is the key enzyme in the final step of anthocyanin biosynthesis, and its function is mainly to transfer glucosyl group which is on UDP-glucose to C3hydroxyl of flower pigment molecules, resulting in formation of the stable anthocyanins in vacuole and the cytoplasm. Up to now, this gene has been cloned from the plants as corn, rice, petunias, and fruits. Many studies have shown that UFGT genes play important roles in the biosynthesis of anthocyanins. In this study, we used purple wheat seed coat as materials, isolated the gene glycosyl transferase TaUFGT involved in anthocyanin biosynthesis, and analyzed this gene biological function and expression pattern.Designed specific primers, using grain seed coat at20days after pollination as materials, we isolated sequences of TaUFGT cDNA by RT-PCR, TaUFGT cDNA length is1374bp, and it encodes457amino acids. This gene had6copies, and named them as TaUFGT-1, TaUFGT-2, TaUFGT-3, TaUFGT-4, TaUFGT-5, TaUFGT-6. Clustering analysis showed TaUFGT-4has a high homology with UFGT gene in barley, belongs to UDP-Glycosyltransferases.In order to confirm the biological function of this gene, we used over-expression vector pBI121-35S:TaUFGT-4and transformed it into the Arabidopsis Atufgt mutant(ugt78d2). The mutant has no pigment accumulation because of the mutation of the UFGT gene. The top section of hypocotyl and cotyledons in the wild type is purple, but the mutant has no purple colour. However the top section of hypocotyl and cotyledons edge in the Arabidopsis Atufgt mutant with pBI121-35S:TaUFGT-4recovered purple colour, indicating that TaUFGT-4can restore the phenotype of mutant.To confirm the gene encoding protein activity, we induced TaUFGT-4protein expression in Pichia pastoris, we get a size of65kDa protein, HPLC analysis showed that enzyme activity of TaUFGT-4protein, but we did not detect the generation of anthocyanin in negative control, suggesting TaUFGT-4protein has glycosyl transferase activity.By qRT-PCR, we detected expression of TaUFGT-4in luozhen No.1seed coat at different stages and different organs, and found that it was expressed in seed coat tissues from10DAP to30DAP, root and leaf tissues. |
