Quantitative trait locus (QTL) analysis was used to study the evolution of genes controlling the timing of flowering in four Brassica genomes that are all extensively replicated. Comparative mapping showed that a chromosomal region from the top of Arabidopsis thaliana chromosome 5 corresponded to three homoeologous copies in each of the diploid species Brassica nigra, B. oleracea, and B. rapa and six copies in the amphidiploid B. juncea. QTLs were detected in two of the three replicated segments in each diploid genome and in three of the six replicated segments in B. juncea. These results indicate that, for the studied trait, multiple QTLs resulting from genome duplication is the rule rather than the exception. Brassica homologues to two candidate genes (CO and FLC) identified from the corresponding A. thaliana region were mapped. CO homologues mapped close to the QTL peaks in eight of nine QTLs, while FLC homologues mapped farther away in those cases where the mapping resolution allowed a comparison. Thus, our data are consistent with the hypothesis that all the major QTLs we detected in the different species of Brassica could be the result of duplicated copies of the same ancestral gene, possibly the ancestor of CO.