Isolation and characterization of the dihydroflavonol 4-reductase gene in the monocotyledonous ornamental Agapanthus praecox ssp. orientalis (Leighton) Leighton

Abstract

Agapanthus praecox ssp. orientalis (Leighton) Leighton, a monocotyledonous ornamental plant belonging to Agapanthaceae, has recently become popular as a potted plant for landscaping and as a cut flower. As a first step toward molecular breeding for flower color alteration in this plant, we isolated and characterized a gene encoding dihydroflavonol 4-reductase (DFR), a pivotal enzyme of the flavonoid biosynthetic pathway. A full-length cDNA clone for DFR was isolated from flower tepals of a cultivar with deep-blue flowers, and its genomic clone, designated ApDFR1 (accession number AB099529 in the GenBank/EMBL/DDBJ databases) was isolated from leaves by a polymerase chain reaction (PCR)-based strategy. Nucleotide sequence analysis revealed that ApDFR1 contains four introns and an open reading frame encoding a polypeptide of 378 amino acid residues. Deduced amino acid sequence shows 59–75% identities with those of previously reported DFR genes. Southern blot analysis showed that there are one or two copies of the DFR gene in the genome of A. praecox ssp. orientalis. ApDFR1 transcripts were detected in young flower tepals, stamens, pistils and bracts, but not in pedicles, scapes and leaves as revealed by reverse transcription-PCR analysis. When ApDFR1 was expressed under the control of the cauliflower mosaic virus 35S promoter in transgenic Petunia hybrida ‘W85’, a dfr-recessive line, some transgenic plants showed drastic flower color alteration from white to red purple. High-performance liquid chromatography analysis showed that colored flower limbs of transgenic plants accumulate anthocyanidins, mainly cyanidin and petunidin. These results indicate that ApDFR1 encodes DFR and is active in a heterologous plant species.