Cloning of the Flavonoid 3&apos;-Hydroxylase Gene of <i>Eustoma grandiflorum</i> (Raf.) Shinn. (<i>EgF3&apos;H</i>) and Complementation of an F3&apos;H-deficient Mutant of <i>Ipomoea nil</i> (L.) Roth. by Heterologous Expression of <i>EgF3&apos;H</i>

Abstract

A full-length cDNA of a putative flavonoid 3'-hydroxylase (F3'H) gene encoding a key enzyme in the production of cyanidin was cloned from a lisianthus (Eustoma grandiflorum) petal. Lisianthus F3'H (EgF3'H) shares 75.1, 73.8, and 68.2% amino acid identity with Arabidopsis thaliana, Ipomoea nil, and Petunia hybrida, respectively. RT-PCR revealed that wild-type lisianthus flowers accumulated higher levels of F3'H mRNA during the early stages of development than in the late stages. The accumulated F3'H transcript levels in leaves were similar to those in flowers in the early stages of development. Overexpression of lisianthus F3'H cDNA altered flower color from red to blue in the I. nil cultivar 'Violet', which lacks a functional F3'H gene. In addition, the transgenic 'Violet' plants accumulated cyanidin and peonidin at similar levels to wild-type I. nil. Taking these findings together, this study demonstrates that EgF3'H functions as a flavonoid 3'-hydroxylase with a role in the synthesis of cyanidin and peonidin pigments.

Cloning and Characterization of a Flavonoid 3′-Hydroxylase Gene from Tea Plant (Camellia sinensis)

“…Five well characterized F3′Hs from other plant species, MdF3′H from Malus × domestica (GenBank ID: ACR14867.1) [ 29 ], PhF3′H from Petunia hybrida (GenBank ID: AAD56282.1) [ 17 ], EgF3′H from Eustoma grandiflorum (GenBank ID: BAP94456.1) [ 30 ], IbF3′H from Ipomoea batatas (GenBank ID: AEH42499.1) [ 31 ] and AtF3′H from Arabidopsis thaliana (GenBank ID: CAB62611.1) [ 18 ] were selected for sequence comparison. Multiple amino acid sequence alignment revealed high homology to these five F3′H sequences.…”

Flavonoid 3',5'-Hydroxylase (<i>F3'5'H</i>) Gene Polymorphisms Co-segregate with Variation in Anthocyanin Composition in the Flower Petals of Lisianthus [<i>Eustoma grandiflorum</i> (Raf.) Shinn.]

“…Young leaves were used for DNA and RNA extraction and petals (not including the throat) were used for RNA extraction at six stages of flower development: 1) bud less than 20-mm long; 2) bud between 20 mm and 30 mm; 3) bud more than 30 mm and tightly closed; 4) bud more than 30 mm and loosely closed; 5) opening flower; and 6) fully opened flower. Extraction was performed and the same materials and stages of flowers were used as described by Takatori et al (2015). Young leaves and petal samples were immediately frozen in liquid nitrogen and stored at −80°C until DNA and RNA extraction.…”

“…Full-length cDNAs of chalcone synthase, chalcone isomerase, flavanone 3hydroxylase, dihydroflavonol 4-reductase, and anthocyanidin synthase were first cloned by Noda et al (2004). cDNAs of F3'5'H, flavonol synthase (FLS), and F3'H were first cloned by Nielsen and Podivinsky (1997), Nielsen et al (2002), and Takatori et al (2015), respectively. Takatori et al (2015) studied F3'H expression in lisianthus strains with various anthocyanin compositions and reported no significant variation in F3'H expression between them.…”

“…cDNAs of F3'5'H, flavonol synthase (FLS), and F3'H were first cloned by Nielsen and Podivinsky (1997), Nielsen et al (2002), and Takatori et al (2015), respectively. Takatori et al (2015) studied F3'H expression in lisianthus strains with various anthocyanin compositions and reported no significant variation in F3'H expression between them.…”

Construction of transgenic <i>Ipomoea obscura</i> that exhibits new reddish leaf and flower colors due to introduction of β-carotene ketolase and hydroxylase genes

“…Modern techniques in plant biotechnology, such as somatic hybrid formation and plant-host transformation followed by its regeneration, are expected to enable further breeding of ornamental Ipomoea species. As for Japanese morning glory (I. nil), many reports have been published on its tissue culture (Jia and Chua 1992;Shimizu et al 2003Shimizu et al , 2005Yoneda and Nakamura 1987), on its transformation (Kikuchi et al 2005;Ono et al 2000;Shibuya et al 2014;Takatori et al 2015;Watanabe et al 2017b) and on its genome editing (Watanabe et al 2017a(Watanabe et al , 2018, whereas few reports have described embryogenic callus formation and plant regeneration in other ornamental Ipomoea species (Ishikuro et al 2014).…”