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The first genetic maps for subterranean clover (Trifolium subterraneum L.) and comparative genomics with T. pratense L. and Medicago truncatula Gaertn. to identify new molecular markers for breeding

Abstract

This study reports on the construction of the first genetic maps of subterranean clover (Trifolium subterraneum L.), a diploid, inbreeding annual pasture legume, and alignment of its linkage groups with those of red clover (T. pratense L.) and Medicago truncatula Gaertn. Transferability of red and white clover (T. repens L.) simple sequence repeat (SSR) markers to subterranean clover was observed. A total of 343 SSR loci were mapped into eight subterranean clover linkage groups, with 6–31 loci per linkage group and 27 loci with similar locations between two distinct F 2 mapping populations. Phenotypic data obtained for flowering time, content of three isoflavonoids (formononetin, genistein and biochanin A), hardseededness, leaf markings, calyx pigmentation and hairiness of stems were analyzed, together with genotypic data. Genomic intervals influencing each trait were assigned to one to three chromosome regions, accounting for 5.5–59.8% of the phenotypic variance. Syntenic relationships were observed among subterranean clover, red clover and Medicago truncatula genomes. Comparisons of loci shared between the three species indicated that at least two chromosomal regions have undergone duplications in the subterranean clover genome. Candidate genes for isoflavone content were identified using M. truncatula as a reference genome. Synteny-based segmentation observed in Brassicaceae chromosomes helped to account for the apparent segmental-based relationship between the clover genomes, particularly within the subterranean clover lines. The proposed segmental nature of clover genome could account for the extensive variation observed between the parental genotypes, while not preventing production of fertile intercrosses.

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Acknowledgments

We are grateful to Hiroshi Hisano (KDRI, currently at Samuel Noble Foundation in USA), Rosemarie Lugg and Nader Aryamanesh (from UWA) for their help in conducting some experiments and data collection. Financial and other support from the Australian Research Council (ARC), Department of Agriculture and Food Western Australia (DAFWA) and Kazusa DNA Research Institute (KDRI) is gratefully acknowledged. Troy Faithful was the recipient of a DAFWA studentship award. Financial support was also obtained from a bequest to UWA by the late Frank Ford.

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Correspondence to Kioumars Ghamkhar.

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Supplementary material 1 (DOCX 34 kb)

11032_2011_9612_MOESM2_ESM.pdf

Fig. S1 Genetic linkage maps of T. subterraneum. The order of LGs has been chosen on the basis of syntenic markers between these maps and the red clover map. QTLs identified for studied traits, as shown in Table 2, are indicated by bars on right-hand side of the linkage groups. Numbers on the left of each group are Kosambi map distances (PDF 22 kb)

11032_2011_9612_MOESM3_ESM.pdf

Fig. S2 Comparative map showing matching of T. subterraneum linkage groups (LGa–LGg) with those of T. pratense (LG1–LG7). T. subterraneum population 92S05 is shown on the left and 92S80 is on the right for each T. pratense LG. Red lines compare positions of shared markers (highlighted in red) between T. pratense and T. subterraneum LGs. Numbers in brackets indicate the number of mapped loci for each LG. Note: the LGe for T. subterraneum population 92S80 did not align with LG5 of T. pratense (PDF 76 kb)

11032_2011_9612_MOESM4_ESM.pdf

Fig. S3 Comparison of SSR marker loci for linkage groups LGf and LGc of T. subterraneum populations 92S05 and 92S80, respectively, and simplified T. pratense linkage group LG6, illustrating synteny between the three LGs. Markers in common with two or more chromosomes are indicated by horizontal bars. Solid rectangles represent the same chromosome segments, with arrows representing the direction of synteny. Numbers represent distance in centimorgans. Note: the entire linkage map of T. pratense LG6 is not shown due to the very high density of molecular markers (PDF 18 kb)

11032_2011_9612_MOESM5_ESM.pdf

Fig. S4 Hypothetical inter-specific shared segments containing isoflavone content genes. Units are cM in clover LGs and Mbp in Medicago truncatula chromosomes (PDF 30 kb)

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Ghamkhar, K., Isobe, S., Nichols, P.G.H. et al. The first genetic maps for subterranean clover (Trifolium subterraneum L.) and comparative genomics with T. pratense L. and Medicago truncatula Gaertn. to identify new molecular markers for breeding. Mol Breeding 30, 213–226 (2012). https://doi.org/10.1007/s11032-011-9612-8

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Keywords

  • Isoflavone
  • Legume
  • Marker-assisted breeding
  • QTL map
  • Segmental duplication
  • Synteny