Crossbreeding of a metallic color carnation and diversification of the peculiar coloration by ion-beam irradiation

In general, carnations (Dianthus caryophyllus) have each of four kinds of anthocyanins acylated by malic acid. A few carnation cultivars are known to display a peculiar dusky color supposedly caused by anthocyanic vacuolar inclusions (AVIs). The hereditary pattern suggests that the peculiar color is controlled by a single recessive factor tightly linked with existence of AVIs containing non-acylated anthocyanins. To diversify the peculiar color carnation, we produced a bluish purple line displaying a highly novel metallic appearance by crossbreeding. By subjecting the line to ion-beam irradiation, we generated metallic reddish purple, metallic crimson and metallic red lines. The major anthocyanin of the metallic bluish purple and reddish purple lines was pelargonidin 3,5-diglucoside, whereas that of the metallic crimson and red lines was pelargonidin 3-glucoside. All four metallic lines did not have transcripts for anthocyanin malyltransferase. Metallic crimson and red lines did not express the acyl-glucose-dependent anthocyanin 5-O-glucosyltransferase gene. In contrast to the dusky color types, metallic lines have highly condensed AVIs and water-clear vacuolar sap in the petal adaxial epidermal cells. Differences in the number of AVIs on the abaxial side were observed within mutants containing the same anthocyanin, thereby affecting their shade and hue. We demonstrated that (1) a factor generating the AVIs is inactivated anthocyanin malyltransferase gene, (2) AVIs in water-clear vacuolar sap in the adaxial epidermal cells generate the novel metallic appearance, and (3) ion beam breeding is a useful tool for increasing metallic colors by changing anthocyanin structure and the level of AVIs.