Accumulation of carotenoids and expression of carotenoid biosynthetic genes and carotenoid cleavage dioxygenase genes during fruit maturation in the juice sacs of 'Tamami,' 'Kiyomi' tangor, and 'Wilking' mandarin

To elucidate the mechanism of the high level of beta-cryptoxanthin (beta-cry) accumulation in 'Tamami,' the relationship between carotenoid accumulation and the expression of carotenoid-related genes during fruit maturation was investigated in the juice sacs of 'Tamami,' a hybrid between 'Kiyomil tangor (Citrus unshiu Marc. x Citrus sinensis Osbeck) and 'Wilking' mandarin (Citrus nobilis Lour. x Citrus deliciosa Ten.), and its parental varieties, 'Kiyomi' tangor and 'Wilking' mandarin. In mature fruit, 'Tamami' predominantly accumulated a high level of beta-cry.'Kiyomi'tangor also accumulated beta-cry. However, the level of beta-cry in'Kiyomi' tangor was lower than that in 'Tamami.' 'Wilking' mandarin accumulated a high level of 9-cis-violaxanthin predominantly. To analyze the expression of carotenoid-related genes, cDNA fragments for their genes were cloned from the three varieties, and the TaqMan MGB probe and a set of primers were designed for the respective genes on the basis of the common sequence of cDNA fragments among the three varieties. In 'Tamami' and 'Wilking' mandarin, the expression of genes that produce beta,beta-xanthophylls (CRPSY, CitPDS, CitZDS, CitLCYb, CitHYb, and CitZEP) increased simultaneously during the massive accumulation of beta,beta-xanthophylls (beta-cry, zeaxanthin, all-trans-violaxanthin, and 9-cis-violaxanthin). In addition, higher levels of upstream synthesis genes (CitPSY, CitPDS, CitZDS, and CitLCYb) and lower levels of downstream synthesis genes (CitHYb and CitZEP) were observed in 'Tamami' than in 'Wilking' mandarin. In 'Kiyomi' tangor, simultaneous increases in the expression of genes that produce beta,beta-xanthophylls were observed in the early maturation stage. During the beta,beta-xanthophyll accumulation, the gene expression of carotenoid cleavage dioxygenases (CitCCD1, CitNCED2, and CitNCED3) increased in the three varieties. The results suggest that the simultaneous increases in gene expression under a balance of high expression of upstream synthesis genes and low expression of downstream synthesis genes led to a high level of beta-cry accumulation in the juice sacs of 'Tamami'.