Comparison of UDP-glucose:flavonoid 3-O-glucosyltransferase (UFGT) gene sequences between white grapes (Vitis vinifera) and their sports with red skin
Introduction
The color of a grape berry is determined by the quantity and composition of anthocyanins in its skin. It is known that red cultivars accumulate anthocyanins in their skins, whereas white cultivars do not synthesize them [1], [2]. Anthocyanin compositions in grape cultivars have been studied by several researchers [2], [3], [4]. Shiraishi and Watanabe [4] determined the anthocyanin compositions of 59 cultivars of red and black grapes and reported that most red cultivars contain mainly derivatives of the anthocyanidins cyanidin and/or peonidin, while black cultivars contain primarily delphinidin, petunidin, and/or malvidin derivatives.
Boss et al. [1], [2] showed that the expression of the UDP-glucose:flavonoid 3-O-glucosyltransferase (UFGT) gene was critical for anthocyanin biosynthesis in the grape berry. Using several tissues of the black cultivar Shiraz, they examined the expression of seven genes involved in anthocyanin biosynthesis and showed that all genes except UFGT were expressed in most tissues. However, UFGT gene expression was only detected in the berry skin, where anthocyanins were present [1]. From the research using the skins of white and red cultivars, they also found that the UFGT gene was expressed in all red cultivars and in none of the white ones [1]. Furthermore, using some bud sports, white- and bronze-skin sports from black-skin cultivars, or red-skin sports from white-skin cultivars, they showed that the UFGT gene expression was only detected in colored grapes that synthesized anthocyanins [2].
From their findings, UFGT is thought to be the key enzyme for anthocyanin biosynthesis in grape. To examine the possibility that the UFGT gene could be absent in the white cultivars, Boss et al. [2] carried out Southern blot analysis on the cultivars using a full-length UFGT cDNA clone as a probe and detected the hybridizing bands in each of the white cultivars [2]. This suggests that the white cultivars possess at least part of the UFGT gene in their genomes. To obtain further information on this gene, using white cultivars and their bud sports with red skin, we isolated UFGT homologues from each cultivar and determined whether the sequences of the genes had differences.
Section snippets
Northern blot analysis
Mature berry skins of Vitis vinifera cv. Italia (white-skin), Ruby Okuyama (a red-skin sport from Italia), Muscat of Alexandria (white-skin), and V. labruscana cv. Kyoho (black-skin) were sampled from vines grown in a field, but those of Flame Muscat (a red-skin sport from Muscat of Alexandria) were taken from vines grown in a greenhouse. The berries (flesh+skin) from the Kyoho vines grown in a greenhouse were collected every two weeks after flowering (WAF) for two to six weeks. Furthermore,
Expression of anthocyanin biosynthetic pathway genes
All of the structural genes for anthocyanin biosynthesis were expressed in a black-skin cultivar Kyoho (Fig. 1). In the white cultivars Italia and Muscat of Alexandria, expression of the genes was much lower than in Kyoho. The expression pattern was also slightly different between Italia and Muscat of Alexandria. Expression of CHS was at a non-detectable level in Italia, and that of PAL was not detected in Muscat of Alexandria. In either cultivar, UFGT gene expression was not detected. Boss et
Acknowledgements
We wish to thank Drs W. Sakamoto and M. Murata of the Research Institute for Bioresources of Okayama University, and Dr R. Nakaune of our Research Center for valuable advice on Northern blot analysis and construction of a cDNA library; J. Kato of the Institute of Fruit Tree Research, Hiroshima Prefectural Agriculture Research Center for providing Flame Muscat; and Dr M. Yamada, A. Sato and H. Iwanami of our Research Center for providing the other grapes. We are also grateful to T. Nakasumi and
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