Two novel inactive alleles of Dihydroflavonol 4-reductase-A (DFR-A) were identified in yellow onion (Allium cepa L.) cultivars and breeding lines from Korea and Japan. Unlike the previously reported inactive yellow DFR-A allele, designated as DFR-A ( TRN ) , in which the 3' portion of the coding sequences was deleted, an allele containing a premature stop codon, DFR-A ( PS ) , was isolated from the majority of cultivars. Co-segregation of DFR-A ( PS ) and color phenotypes in the F(2) population from a cross between yellow and red parents showed that inactivation of DFR-A was responsible for lack of anthocyanin in these yellow onions. In addition, RT-PCR analysis of F(2) population showed that the transcription level of the DFR-A ( PS ) allele was significantly reduced owing to non-sense-mediated mRNA decay. A 20-bp deletion of a simple sequence repeat in the promoter region of the DFR-A ( PS ) allele was used to develop a simple PCR-based molecular marker for selection of the DFR-A ( PS ) allele. All genotypes of 138 F(2) individuals were clearly distinguished by this molecular marker. In addition to the DFR-A ( PS ) allele, another DFR-A allele, DFR-A ( DEL ) , was identified in some cultivars. In case of the DFR-A ( DEL ) allele, no PCR products were amplified throughout DFR-A sequences including promoter regions, suggesting deletion of the entire DFR-A gene. Co-segregation of the absence of DFR-A and color phenotypes was confirmed in another F(2) population. Furthermore, RT-PCR results showed that no DFR-A transcript was detected in any yellow F(2) individuals.