We sequenced five BAC clones of Brassica oleracea doubled haploid 'Early Big' broccoli containing major genes in the aliphatic glucosinolate pathway, and comparatively analyzed them with similar sequences in A. thaliana and B. rapa. Additionally, we included in the analysis published sequences from three other B. oleracea BAC clones and a contig of this species corresponding to segments in A. thaliana chromosomes IV and V. A total of 2,946 kb of B. oleracea, 1,069 kb of B. rapa sequence and 2,607 kb of A. thaliana sequence were compared and analyzed. We found conserved collinearity for gene order and content restricted to specific chromosomal segments, but breaks in collinearity were frequent resulting in gene absence likely not due to gene loss but rearrangements. B. oleracea has the lowest gene density of the three species, followed by B. rapa. The genome expansion of the Brassica species, B. oleracea in particular, is due to larger introns and gene spacers resulting from frequent insertion of DNA transposons and retrotransposons. These findings are discussed in relation to the possible origin and evolution of the Brassica genomes.