Gene characterization, analysis of expression and in vitro synthesis of dihydroflavonol 4-reductase from [Citrus sinensis (L.) Osbeck]

Publication Overview
TitleGene characterization, analysis of expression and in vitro synthesis of dihydroflavonol 4-reductase from [Citrus sinensis (L.) Osbeck]
AuthorsLo Piero A, Puglisi I, Petrone G
TypeJournal Article
Journal NamePhytochemistry
Volume67
Year2006
Page(s)684-695
CitationLo Piero A, Puglisi I, Petrone G. Gene characterization, analysis of expression and in vitro synthesis of dihydroflavonol 4-reductase from [Citrus sinensis (L.) Osbeck]. Phytochemistry. 2006; 67:684-695.

Abstract

Dihydroflavonol 4-reductase (DFR, EC 1.1.1.219) catalyzes the reduction of dihydroflavonols to leucoanthocyanins, a key "late" step in the biosynthesis of anthocyanins. In this study we showed that a strong reduction in DFR expression occurs in the non-red orange cultivar (Navel and Ovale) compared to that of the red orange (Tarocco) suggesting that the enzyme could be involved in the lack of production of anthocyanins. Therefore, we isolated and compared the cDNAs, the genomic clones, as well as the promoter regions of blood and blond orange dfrs. Our data revealed that the cDNA sequences of pigmented and non-pigmented orange DFRs were 100% homologous and contained a 1017 bp open reading frame which encodes a protein of 338 amino acid residues, corresponding to a molecular mass of 38010.76 Da, with a theoretical pI of 5.96. Moreover, we found that there were no significant differences in non-coding regions (introns and 5' upstream region) of dfr sequences. Southern blot analysis of genomic DNA indicated that dfr was present as a single copy gene in both cultivars. From these findings the low expression level of blond orange dfr, which might play a role in the phenotypic change from blood to blond orange, is thought to be the result of a likely mutation in a regulatory gene controlling the expression of dfr. In addition, here we reported the successful expression of orange DFR cDNAs leading to an active DFR enzyme which converts dihydroquercetin to leucoanthocyanidin, thus confirming the involvement of the isolated genes in the biosynthesis of anthocyanins. Moreover, as far as we know, this is the first report concerning the in vitro expression of DFR from fruit flesh whose biochemical properties might be very different from those of other plant organ DFRs.
Features
This publication contains information about 15 features:
Feature NameUniquenameType
AP1AP1genetic_marker
AP2AP2genetic_marker
DFR50UTRDFR50UTRgenetic_marker
DFR-GSP1- revDFR-GSP1- revgenetic_marker
DFR-GSP1/2-revDFR-GSP1/2-revgenetic_marker
DFR-GSP2/2-revDFR-GSP2/2-revgenetic_marker
DFR-GSP2-revDFR-GSP2-revgenetic_marker
DFRPOL2DFRPOL2genetic_marker
DFR-probeDFR-probegenetic_marker
for-Citrus-DFRfor-Citrus-DFRgenetic_marker
rev-Citrus-DFRrev-Citrus-DFRgenetic_marker
AY498567AY498567.1region
AY519363AY519363.1region
DQ084723DQ084723.1region
DQ084722DQ084722.1region
Stocks
This publication contains information about 3 stocks:
Stock NameUniquenameType
NavelNavelaccession
TaroccoTaroccoaccession
ValenciaValenciaaccession
Properties
Additional details for this publication include:
Property NameValue
URLhttp://dx.doi.org/10.1016/j.phytochem.2006.01.025
Published Location|||
Language Abbreng
Publication Model[electronic resource].
Publication TypeJournal Article
Publication Date2006
KeywordsCitrus sinensis, Rutaceae, cultivars, oranges, fruits (plant anatomy), plant extracts, methyltransferases, complementary DNA, promoter regions, sequence analysis, open reading frames, introns, gene expression regulation, anthocyanins, plant pigments, biosynthesis, Southern blotting, amino acid sequences, nucleotide sequences, dihydroflavonol 4-reductase, 5'-noncoding region, molecular sequence data