Study Of Color Mutation Of Grape Berry Skin In Black-’Brazil’ And Its Molecular Mechanism

In order to study the reason of color mutation of grape berry skin, from red-’Benitaka’ to black-’Brazil’,we analyzed the genotypes of the VvmybAl loci in white-’Italia’, black-’Brazil’and red-’Benitaka’,also the levels of VvmybAl and UFGT expression in berry skins of different developmental stages were determined. We cloned the promoter region and coding sequence of VvmybAl from these three materials.Their functions were analysised by transient expression system.The main results were listed as follows:1.22pairs of polymorphic SSR primers were used to analyze the grapevine cultivars and judge their genetic relationships. In this study,’Italia’,’Brazil’and’Benitaka’didn’t show any SSR polymorphism at any of the22SSR loci tested. These results revealed that the genotypes of ’Brazil’ and ’Benitaka’ were identical to that of’Italia’, and both red-’Benitaka’ and black-’Brazil’ are bud sports of white-’Italia’.2. We found white-’Italia’contained only VvmybAla (1559bp),’Benitaka’ and ’Brazil’were heterozygous for the alleies VvmybA1a and VvmybA1BEN (926bp) at VvmybA1loci. QRT-PCR analysis was carried out to compare the relative expression levels of VvmybAl and UFGT in these three cultivars. Expression of VvmybAl and UFGT was almost undetectable in white-’Italia’,and their expression mode were at equal pace in colored cultivars, which could be detected when the berries began to color. The level of VvmybAl and UFGT expression in’Brazil’was significantly higher than that of’Benitaka’.3. We cloned the promoter region and full CDS of VvmybAl from three cultivars, and compared that with known sequences by BioXM2.6. We also used Plant Care and Place to analysis the promoter region. There was only1base substitution between’Brazil’and ’Benitaka’ within their promoter regions, according to Plant Care we found this position was at3-AF1binding site which was light responsive element. Three cultivars’promoter regions and full-length coding regions of VvmybAl were integrated into PYH4215without35S promoter. Agrobacterium-mediated transformation was adopted to introduce this chimeric gene into somatic embros. Upon the introduction of VvmybA1from Brazil and Benitaka, red cells were induced in the embryos, whereas the introduction of VvmybAl from Italia failed to do so. That meaned the novel VvmybA1BEN alleles from Brazil and Benitaka were shown to have ability to restore VvmybA1transcripts.