Expression of the dihydroflavonol reductase gene in an anthocyanin-free barley mutant.

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The barley gene encoding dihydroflavonol-4-reductase (DFR) was delivered by micoprojectile bombardment into leaf sheath tissue of the anthocyanin-free barley mutant ant 18-162, a mutant which lacks DFR activity-probably because of a missense mutation in the structural gene for DFR. The delivered gene complemented the mutation, as evidenced by the synthesis of anthocyanin in individual leaf sheath cells of the bombarded tissues. Pigment synthesis appeared two days after gene delivery and both the number of pigmented cells and the intensity of pigmentation increased over the following days. Depending on the physiological condition of the host plants, up to 15 pigmented cells per 10 tissue segments were detected. These results demonstrate that the Ant 18 gene of barley encodes dihydroflavonol-4-reductase. A series of gene constructs encoding DFR were expressed in the anthocyanin-free mutant tissue. The genomic clone complemented the mutation whereas an equivalent plasmid with all introns deleted did not. The highest number of pigmented cells was obtained using plasmids containing the DFR-coding sequence interrupted by intron 1 of the genomic clone, indicating that the presence of an intron stabilizes the DFR message.