Background: There are three basic Brassica genomes (A, B, and C) and three parallel sets of subgenomes distinguished in the diploid Brassica (i.e.: B. rapa, ArAr; B. nigra, BniBni; B. oleracea, CoCo) and the derived allotetraploid species (i.e.: B. juncea, AjAjBjBj; B. napus, AnAnCnCn; B. carinata, BcBcCcCc). To understand subgenome differentiation in B. juncea in comparison to other A genome-carrying Brassica species (B. rapa and B. napus), we constructed a dense genetic linkage map of B. juncea, and conducted population genetic analysis on diverse lines of the three A-genome carrying Brassica species using a genotyping-by-sequencing approach (DArT-seq). Results: A dense genetic linkage map of B. juncea was constructed using an F2 population derived from Sichuan Yellow/Purple Mustard. The map included 3329 DArT-seq markers on 18 linkage groups and covered 1579 cM with an average density of two markers per cM. Based on this map and the alignment of the marker sequences with the physical genome of Arabidopsis thaliana, we observed strong co-linearity of the ancestral blocks among the different A subgenomes but also considerable block variation. Comparative analyses at the level of genome sequences of B. rapa and B. napus, and marker sequence anchored on the genetic map of B. juncea, revealed a total of 30 potential inversion events across large segments and 20 potential translocation events among the three A subgenomes. Population genetic analysis on 26 accessions of the three A genome-carrying Brassica species showed that the highest genetic distance were estimated when comparing Aj-An than between An-Ar and Aj-Ar subgenome pairs. Conclusions: The development of the dense genetic linkage map of B. juncea with informative DArT-seq marker sequences and availability of the reference sequences of the Ar, and AnCn genomes allowed us to compare the A subgenome structure of B. juncea (Aj). Our results suggest that strong co-linearity exists among the three A Brassica genomes (Ar, An and Aj) but with apparent subgenomic variation. Population genetic analysis on three A-genome carrying Brassica species support the idea that B. juncea has distinct genomic diversity, and/or evolved from a different A genome progenitor of B. napus.