Abstract
Background: Molecular markers and genetic linkage maps are pre-requisites for molecular breeding, but no genetic map is currently available for subterranean clover (Trifolium subterraneum L.), a diploid and inbreeding pasture legume. A total of 2,646 microsatellite or simple sequence repeat (SSR) markers either available in the public domain or unpublished obtained from Kazusa DNA Research Institute, were screened on four genotypes (Denmark, DGI007, Woogenellup and Daliak) that are parents of two F2 mapping populations. Results: A total of 343 SSR loci could be mapped into 8 linkage groups with 6–31 loci per linkage group and 27 SSRs shared between the two populations. Phenotyping data obtained for flowering time, isoflavonoid content (formononetin, genistein, biochanin A), hardseedness, leaf marks, calyx pigmentation and hairiness of stem were analyzed together with genotyping data. For each trait, 1–3 QTLs were identified; the phenotypic variation explained by these QTLs was in the range of 5.5–59.8%. In addition, alignment of linkage groups was possible with the genetic maps of red clover (Trifolium pratense L.), white clover (Trifolium repens L.)and Medicago truncatula Gaertn. using shared loci. Segmental duplications were identified within subterranean clover genome and between the legume species. Candidate genes for isoflavone content were identified. Conclusions: The present study reports on comparative genomics among three clover species as well as Medicago truncatula following the construction of the first genetic map for subterranean clover. It demonstrates the utility of a comparative genomics approach for molecular mapping of QTL controlling seed hardness, flowering time and specifically isoflavone content traits, as well as establishing relationships between subterranean clover and genomes of red clover, white clover and model legume M. truncatula. The comparative mapping carried out demonstrated the value of the map to identify candidate genes controlling important traits such as isoflavone content.