Inheritance of Flower Pigment in Crosses between Cyclamen Cultivars and Cyclamen purpurascens

Although interspecific hybrids between cyclamen (Cyclamen persicum) cultivars and C. purpurascens were produced by using the ovule culture, mode of inheritance of flower pigment remains obscure. Therefore, inheritance of anthocyanin in the petals in the crosses between diploid cyclamen cultivars and C. purpurascens was investigated in the present study. In every cross combination, including the crosses by using acyanic cyclamen cultivars without the eye, all the F1 progenies had a purple or reddish-purple eye in the petals, showing the eye expression was a dominant characteristic. All the F1 progenies contained anthocyanins in the petals, whereas some progenies in the crosses by using a yellow-flowered cultivar contained very small amount of anthocyanin. The main flower pigment of C. purpurascens was malvidin 3,5-diglucoside. All the F1 progenies also contained 3,5diglucoside type anthocyanins, such as malvidin 3,5-diglucoside, peonidin 3,5-diglucoside and cyanidin 3,5-diglucoside, in the petals as the main pigments even in the crosses by using acyanic cultivars or cyanic cultivars containing malvidin 3-glucoside or peonidin 3-neohesperidoside in the petals as the main pigment. These results suggest that the expression of 5-glucosyltransferase in anthocyanin synthesis in the petals was dominant even in the crosses between cyclamen cultivars and C. purpurascens. INTRODUCTION Among the genus Cyclamen consisting of 22 species (Grey-Wilson, 2003), only C. persicum has been used as the major commercial plants. The other Cyclamen species, however, have desirable characteristics like sweet fragrance in C. purpurascens. Although intespecific hybridization between C. persicum and the other species had been difficult because of the strong cross-incompatibility (Legro, 1959; Grey-Wilson, 2003), Ishizaka and Uematsu (1992) overcame the difficulty with the ovule culture system. The system was also used successfully for obtaining interspecific hybrids between C. persicum cultivars and C. purpurascens (Ishizaka and Uematsu, 1995a; Ewald, 1996; Takamura et al., 2002), and the hybrids had sweet fragrance in their flower like C. purpurascens (Ishizaka and Uematsu, 1995a; Ewald, 1996). Flower color is another important characteristic for ornamental plants so that cultivars with desirable color and fragrance are demanded. While there are a few reports on the inheritance in crosses among cyclamen cultivars (Van Bragt, 1962; Takamura et al., 1995, 2000a,b,c), no report on the inheritance in interspecific crosses in the genus Cyclamen is available. Therefore, we investigated inheritance of anthocyanin in the petals in crosses between diploid cyclamen cultivars and C. purpurascens. Proc. IX Intl. Symp. on Flower Bulbs Eds.: H. Okubo, W.B. Miller and G.A. Chastagner Acta Hort. 673, ISHS 2005 438 MATERIALS AND METHODS Diploid cyclamen (C. persicum) cultivars were crossed with C. purpurascens and their F1 progenies were obtained through the ovule culture following Ishizaka and Uematsu (1995a). In diploid cyclamen cultivars, yellow-flowered ‘Golden Boy’, white-flowered ‘Mirabelle’, red-flowered ‘Largo’ and ‘Strauss’, reddish-purple-flowered ‘Cochineal’, and purple-flowered ‘Wase Murasaki’ were used. The petals of F1 progenies were collected, and the flower colors were recorded. Anthocyanins in petals of the F1 progenies and some mother plants were investigated by using high performance liquid chromatography (HPLC). For the HPLC analysis, only slips (petals without their eye parts) were used. The slips were dried, and then soaked in 5% HCOOH methanolic solution at 5°C overnight. The extract was filled up to 5 ml. The filtered petal extract was injected to HPLC, in which chromatography systems were developed with LC-10AT pumps (Shimadzu Co. Ltd., Japan) equipped with Cosmosil 5C18 AR-II columns (4.6 mm I.D. x 50 mm + 4.6 mm I.D. x 250 mm; Nakarai Tesque Co. Ltd., Japan) and an SPD-10AV detector (Shimadzu Co. Ltd.) set at 530 nm. The column temperature was maintained at 40°C. Liner gradient elution for 40 min from 25 to 40 or 41% solvent B (1.5% H3PO4, 20% HCOOH and 25% CH3CN in H2O) in solvent A (1.5% H3PO4 in H2O) were employed as the solvent system, and the flow rate of 0.8 ml was maintained. RESULTS AND DISCUSSION All the F1 progenies in every cross combination, including the crosses by using acyanic cyclamen cultivars without cyanic eye, had a purple or reddish-purple eye in their petals (data not shown). The results suggest that the eye expression was dominant in the crosses as Takamura et al. (2000a) indicated that the eye expression in intraspecific cyclamen crosses was dominant. Van Bragt (1962) and Takamura et al. (2000b) reported that anthocyanin expression in slip parts of cyclamen petals was a dominant characteristic. All the F1 progenies in the present study contained anthocyanins in their slips, suggesting that the anthocyanin expression was also dominant in the interspecific crosses (Table 1). All the F1 progenies also contained 3,5-diglucoside type anthocyanins, such as malvidin 3,5-diglucoside (Mv3,5dG), peonidin 3,5-diglucoside (Pn3,5dG) and cyanidin 3,5-diglucoside (Cy3,5dG), in the slips as the main pigments even in the crosses by using acyanic cultivars or cyanic cultivars of which main anthocyanins in the slips were malvidin 3-glucoside or peonidin 3-neohesperidoside. Van Bragt (1962) and Takamura et al. (2001) suggested that synthesis of anthocyanidin 3,5-diglucoside was dominant in cyclamen cultivars. Results of the present study suggest that the expression of 5-glucosyltransferase in anthocyanin synthesis in the slips may be also dominant in the interspecific crosses. Since the main flower pigment of C. purpurascens was Mv3,5dG, genes in C. purpurascens should contribute to the expression. In ‘Golden Boy’ x C. purpurascens crosses some F1 progenies contained a little anthocyanin in their slips, whereas the other progenies accumulated relatively much anthocyanin. The differences in anthocyanin accumulation in the progenies caused the difference in flower color expression (Fig. 1): The slips of the progenies with a little anthocyanin were very pale pink. In ‘Largo’ x C. purpurascens crosses all the F1 progenies contained Cy3,5dG, Mv3,5dG and Cy3,5dG in their slips (Table 1). However, the primary anthocyanin was Mv3,5dG in some F1 progenies, and the others contained Pn3,5dG in their slips as the primary anthocyanin (data not shown). A chimeric plant with both pink and purple flowers were found in progenies obtained by ‘Strauss’ x C. purpurascens crosses. The main anthocyanin of the pink flowers differed from that of the purple flowers (Fig. 2). The factors that caused the segregation in anthocyanin accumulation and chimeric plant production in the cross combinations remains obscure, and further research on the factors is desirable Results of the present study should contribute to clarify inheritance of flower pigment in interspecific crosses between diploid cyclamen cultivars and C. purpurascens. The F1 progenies in the crosses were, however, sterile (data not shown). Ishizaka and