Molecular Genetics and Genomics

, Volume 280, Issue 4, pp 293–304 | Cite as

Identification of homologous, homoeologous and paralogous sequence variants in an outbreeding allopolyploid species based on comparison with progenitor taxa

  • Melanie L. Hand
  • Rebecca C. Ponting
  • Michelle C. Drayton
  • Kahlil A. Lawless
  • Noel O. I. Cogan
  • E. Charles Brummer
  • Timothy I. Sawbridge
  • German C. Spangenberg
  • Kevin F. Smith
  • John W. Forster
Original Paper


The combination of homologous, homoeologous and paralogous classes of sequence variation presents major challenges for SNP discovery in outbreeding allopolyploid species. Previous in vitro gene-associated SNP discovery studies in the allotetraploid forage legume white clover (Trifolium repens L.) were vulnerable to such effects, leading to prohibitive levels of attrition during SNP validation. Identification of T. occidentale and T. pallescens as the putative diploid progenitors of white clover has permitted discrimination of the different sequence variant categories. Amplicons from selected abiotic stress tolerance-related genes were obtained using mapping family parents and individuals from each diploid species. Following cloning, progenitor comparison allowed tentative assignment of individual haplotypes to one or other sub-genome, as well as to gene copies within sub-genomes. A high degree of coincidence and identity between SNPs and HSVs was observed. Close similarity was observed between the genome of T. occidentale and one white clover sub-genome, but the affinity between T. pallescens and the other sub-genome was weaker, suggesting that a currently uncharacterised taxon may be the true second progenitor. Selected validated SNPs were attributed to individual sub-genomes by assignment to and naming of homoeologous linkage groups, providing the basis for improved genetic trait-dissection studies. The approach described in this study is broadly applicable to a range of allopolyploid taxa of equivocal ancestry.


Forage legume White clover Single nucleotide polymorphism Haplotype Candidate gene Functional variation 



This work was supported by funding from the Victorian Department of Primary Industries, Dairy Australia Ltd., the Geoffrey Gardiner Dairy Foundation, Meat and Livestock Australia Ltd. and the Molecular Plant Breeding Cooperative Research Centre (MPB CRC). The authors thank Dr. Nick Ellison (AgResearch New Zealand) for provision of T. pallescens genomic DNA and both Dr. Ross Chapman and Prof. Michael Hayward for careful critical reading of the manuscript.

Supplementary material

438_2008_365_MOESM1_ESM.doc (90 kb)
MOESM1 Summary information for GenBank submissions of sub-genome specific DNA sequences derived from template genes in this study. Definitions, accession numbers, lengths, nucleotide content, feature descriptions and text versions of sequences are provided. Consensus sequences for multiple haplotypes are provided, with sub-genome specific SNPs coded as redundant bases (M = A or C; R = A or G; W = A or T; S = C or G; Y = C or T; K = G or T; V = A, C or G; H = A, C or T; D = A, G or T; b = C, G or T; N = A, T, C or G) (DOC 89 kb)
438_2008_365_MOESM2_ESM.ppt (40 kb)
MOESM2 Distribution of SNP loci within the structure of the TrDHNb dehydrin gene. The two exons and a single intron are indicated as black and grey boxes, respectively. Positions of LAPs used to generate genomic amplicons are indicated, while arrows denote SNPs in the exons (12) and intron (7) (PPT 39 kb)
438_2008_365_MOESM3_ESM.doc (126 kb)
MOESM3 Full information for levels of nucleotide identity between T. occidentale and T. pallescens reference sequences and O and P' sub-genome sequences. Average values for the consensus sequences of each white clover mapping family parental genotype are presented for each gene (DOC 125 kb)


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Copyright information

© Springer-Verlag 2008

Authors and Affiliations

  • Melanie L. Hand
    • 1
    • 4
  • Rebecca C. Ponting
    • 1
    • 4
  • Michelle C. Drayton
    • 1
    • 4
  • Kahlil A. Lawless
    • 1
    • 4
  • Noel O. I. Cogan
    • 1
    • 4
  • E. Charles Brummer
    • 2
  • Timothy I. Sawbridge
    • 1
    • 4
  • German C. Spangenberg
    • 1
    • 4
  • Kevin F. Smith
    • 3
    • 4
  • John W. Forster
    • 1
    • 4
  1. 1.Department of Primary Industries, Biosciences Research DivisionVictorian AgriBiosciences CentreBundooraAustralia
  2. 2.Centre for Applied Genetic Technologies, Crop and Soil Science DepartmentUniversity of GeorgiaAthensUSA
  3. 3.Department of Primary Industries, Biosciences Research DivisionHamilton CentreHamiltonAustralia
  4. 4.Molecular Plant Breeding Cooperative Research CentreVictorian AgriBiosciences CentreBundooraAustralia

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