Clonging DFR And UFGT Gene From Mango (Mangifera Indica)and Effect On Fruit Coloring

Mangifera indica L., commonly known as Mango, is an evergreen arbor plant of Mangifera L.,it belongs to the Anacardiaceae and is one of the most famous fruit tree species in tropical and subtropical regions, it has an important economic value and nutritional value. The color of Mango fruit directly affects the mental state of the consumer so as to determine the value of the goods. Anthocyanins are flavonoids hormone, which has a significant impact on fruit color. Dihydroflavonol4-reductase (DFR) is an important enzyme in late anthocyanin metabolism, is the key regulatory point to determine the anthocyanin from colorless to colored. Flavonoid glycosyltransferase (UFGT) can catalyze colorless unstable anthocyanin to a stable anthocyanin, mechanism is the sugar-base connected to the instability of anthocyanin, is the last enzyme genes of anthocyanin synthesis process. In this study, several different colored of ripe period of mango varieties was used as materials, using RACE method to extend the full-length cDNA sequences of DFR and UFGT gene, and has carried on the research of bioinformatics and expression characteristics. We can determine the number of introns according to the design of specific primers to amplify full-length sequence in the base group, at the same time, we can determine the expression of these two genes in different mango varieties by fluorescence quantitative PCR, and determine the impact of DFR and UFGT gene on mango fruit coloration combined with physiological and biochemical analysis of the different stages of development. We hope to provide some experimental support for the study of part of the mechanism of the mango fruit coloration through the study of these two genes, at the same time, to provide a theoretical basis for the regulation of the production of mango fruit color and mature period. The main results of the research are as follows:1、In this study, the ’Royal’ mango DFR gene was cloned for the first time, and its full-length of cDNA is1260bp consisted of987bp open reading frame and encoding328amino acids. Further amplified the genomic DNA, the total length is3022bp, analysis showed that it contained five introns and expression conserved in have been reported plants. Phylogenetic analysis showed that mango DFR protein has a close genetic relationship with anthurium, wheat, barley and other plants. Through the analysis of the expression of DFR gene in seven colored peel of mature period of red ’Royal’mango, Yellow’Jinhuang’ mango, green’Gui seven’ mango, found expressed higher levels in green mango peel, while lower amounts expressed in yellow rind, red ’Royal’mango is not expressed, suggesting that further study for DFR regulation of anthocyanin synthesis is need.2、In this study, we cloned UFGT gene from mango for the first time. According to the known UFGT gene fragment, we cloned the full-length cDNA of UFGT with3’ RACE and5’ RACE technology. Using specific primers amplified the full-length sequence of genomic DNA of the gene and found this gene contains only one intron. The full-length cDNA sequences of this gene was constructed into the plant expression vector pET32a, we get a fusion protein about550KDa size by1mmol/L IPTG induction expression.