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Species-Level Phylogeny and Polyploid Relationships in Hordeum (Poaceae) Inferred by Next-Generation Sequencing and In Silico Cloning of Multiple Nuclear Loci.

Brassac J, Blattner FR - Syst. Biol. (2015)

Bottom Line: For diploid taxa, a Bayesian concordance analysis and a coalescent-based dated species tree was inferred from all gene trees.The resulting multilocus phylogeny reveals for the first time species phylogeny and progenitor-derivative relationships of all di- and polyploid Hordeum taxa within a single analysis.Our study proves that it is possible to obtain a multilocus species-level phylogeny for di- and polyploid taxa by combining PCR with next-generation sequencing, without cloning and without creating a heavy load of sequence data.

View Article: PubMed Central - PubMed

Affiliation: Institute of Plant Genetics and Crop Plant Research (IPK), D-06466 Gatersleben, Germany; brassac@ipk-gatersleben.de.

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Scheme summarizing phylogenetic relationships of species and cytotypes in the genus Hordeum based on 12 loci and one chloroplast region. Diploid taxa were drawn directly at the tree obtained from MSC analysis (Fig. 2) modified to reflect differences with BI analysis based on 13 (Fig. 1) and 26 loci (Supplementary Fig. S17) and position of extinct taxa inferred from the BI tree including diploid and polyploids (Supplementary Fig. S19). Tetra- and hexaploids were connected to their inferred progenitors. Double lines indicate autopolyploid origin, dashed lines mark uncertainties and colors refer to the direction of crosses resulting in allopolyploid taxa (blue for paternal, red for maternal parent, black for both observed) based on the chloroplast data (Supplementary Fig. S14). Asterisks indicate incongruent relationships of polyploids recovered by chloroplast sequences (Jakob and Blattner 2006) compared to nuclear sequences that probably indicate polyphyletic origins.
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Figure 3: Scheme summarizing phylogenetic relationships of species and cytotypes in the genus Hordeum based on 12 loci and one chloroplast region. Diploid taxa were drawn directly at the tree obtained from MSC analysis (Fig. 2) modified to reflect differences with BI analysis based on 13 (Fig. 1) and 26 loci (Supplementary Fig. S17) and position of extinct taxa inferred from the BI tree including diploid and polyploids (Supplementary Fig. S19). Tetra- and hexaploids were connected to their inferred progenitors. Double lines indicate autopolyploid origin, dashed lines mark uncertainties and colors refer to the direction of crosses resulting in allopolyploid taxa (blue for paternal, red for maternal parent, black for both observed) based on the chloroplast data (Supplementary Fig. S14). Asterisks indicate incongruent relationships of polyploids recovered by chloroplast sequences (Jakob and Blattner 2006) compared to nuclear sequences that probably indicate polyphyletic origins.

Mentions: To identify the progenitors of the polyploid species, a nuclear loci supermatrix including the sequences derived from diploid and polyploid taxa was phylogenetically analyzed with BI (Supplementary Fig. S19). The positions of the different homoeologues of polyploids in relation to their closest relatives derived from diploids were used to infer the lineages contributing to polyploids. If sequences from a polyploid lineage grouped within different diploids, this was interpreted as an indication for allopolyploidy, while autopolyploidy was inferred if all sequences of a polyploid were in a clade with a single diploid. Clades consisting solely of polyploid-derived sequences were interpreted as indication of extinct progenitor lineages (Blattner 2004; Jakob and Blattner 2010; Brassac et al. 2012). The results of this analysis were summarized into a scheme where polyploids were integrated in the modified diploid species tree (Fig. 3). The MSC topology was modified to take into account the incongruences between the different methods and to integrate the inferred extinct lineages. The polyploid relationships could mostly be identified with confidence. The wide genetic variety found in some species probably indicates multiple origins of such polyploids. Hordeum parodii, a hexaploid species, as well as H. tetraploidum, together with H. fuegianum one of its potential tetraploid progenitors , appeared to be polyphyletic involving the two closely related diploid species H. chilense and H. flexuosum. The partially autopolyploid taxon H. brevisubulatum, containing both auto- and allopolyploid individuals (Brassac et al. 2012), was treated differently. The high diversity of the copies recovered for the different individuals and the difficulty to assign the parental species/individuals across loci made it difficult to create phased haplotypes. Only one tetraploid (PI401387) appeared autopolyploid with gene copies clustering essentially with the species' diploid cytotypes while sequences of other individuals clustered with species within the Asian Hordeum clade (H. roshevitzii and H. bogdanii). The position of many sequences (for example Bre_PI401376_C at BLZ1, Supplementary Fig. S2) retrieved for the polyploid taxa indicates ongoing intergenomic recombination as already suggested by the high proportion of chimerical sequences in cloned TOPO6 sequences (Brassac et al. 2012). Sequencing and analysis of the individual H00312 revealed it to be probably mislabeled and it was then excluded from our conclusions (see Section “Discussion”).


Species-Level Phylogeny and Polyploid Relationships in Hordeum (Poaceae) Inferred by Next-Generation Sequencing and In Silico Cloning of Multiple Nuclear Loci.

Brassac J, Blattner FR - Syst. Biol. (2015)

Scheme summarizing phylogenetic relationships of species and cytotypes in the genus Hordeum based on 12 loci and one chloroplast region. Diploid taxa were drawn directly at the tree obtained from MSC analysis (Fig. 2) modified to reflect differences with BI analysis based on 13 (Fig. 1) and 26 loci (Supplementary Fig. S17) and position of extinct taxa inferred from the BI tree including diploid and polyploids (Supplementary Fig. S19). Tetra- and hexaploids were connected to their inferred progenitors. Double lines indicate autopolyploid origin, dashed lines mark uncertainties and colors refer to the direction of crosses resulting in allopolyploid taxa (blue for paternal, red for maternal parent, black for both observed) based on the chloroplast data (Supplementary Fig. S14). Asterisks indicate incongruent relationships of polyploids recovered by chloroplast sequences (Jakob and Blattner 2006) compared to nuclear sequences that probably indicate polyphyletic origins.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

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Figure 3: Scheme summarizing phylogenetic relationships of species and cytotypes in the genus Hordeum based on 12 loci and one chloroplast region. Diploid taxa were drawn directly at the tree obtained from MSC analysis (Fig. 2) modified to reflect differences with BI analysis based on 13 (Fig. 1) and 26 loci (Supplementary Fig. S17) and position of extinct taxa inferred from the BI tree including diploid and polyploids (Supplementary Fig. S19). Tetra- and hexaploids were connected to their inferred progenitors. Double lines indicate autopolyploid origin, dashed lines mark uncertainties and colors refer to the direction of crosses resulting in allopolyploid taxa (blue for paternal, red for maternal parent, black for both observed) based on the chloroplast data (Supplementary Fig. S14). Asterisks indicate incongruent relationships of polyploids recovered by chloroplast sequences (Jakob and Blattner 2006) compared to nuclear sequences that probably indicate polyphyletic origins.
Mentions: To identify the progenitors of the polyploid species, a nuclear loci supermatrix including the sequences derived from diploid and polyploid taxa was phylogenetically analyzed with BI (Supplementary Fig. S19). The positions of the different homoeologues of polyploids in relation to their closest relatives derived from diploids were used to infer the lineages contributing to polyploids. If sequences from a polyploid lineage grouped within different diploids, this was interpreted as an indication for allopolyploidy, while autopolyploidy was inferred if all sequences of a polyploid were in a clade with a single diploid. Clades consisting solely of polyploid-derived sequences were interpreted as indication of extinct progenitor lineages (Blattner 2004; Jakob and Blattner 2010; Brassac et al. 2012). The results of this analysis were summarized into a scheme where polyploids were integrated in the modified diploid species tree (Fig. 3). The MSC topology was modified to take into account the incongruences between the different methods and to integrate the inferred extinct lineages. The polyploid relationships could mostly be identified with confidence. The wide genetic variety found in some species probably indicates multiple origins of such polyploids. Hordeum parodii, a hexaploid species, as well as H. tetraploidum, together with H. fuegianum one of its potential tetraploid progenitors , appeared to be polyphyletic involving the two closely related diploid species H. chilense and H. flexuosum. The partially autopolyploid taxon H. brevisubulatum, containing both auto- and allopolyploid individuals (Brassac et al. 2012), was treated differently. The high diversity of the copies recovered for the different individuals and the difficulty to assign the parental species/individuals across loci made it difficult to create phased haplotypes. Only one tetraploid (PI401387) appeared autopolyploid with gene copies clustering essentially with the species' diploid cytotypes while sequences of other individuals clustered with species within the Asian Hordeum clade (H. roshevitzii and H. bogdanii). The position of many sequences (for example Bre_PI401376_C at BLZ1, Supplementary Fig. S2) retrieved for the polyploid taxa indicates ongoing intergenomic recombination as already suggested by the high proportion of chimerical sequences in cloned TOPO6 sequences (Brassac et al. 2012). Sequencing and analysis of the individual H00312 revealed it to be probably mislabeled and it was then excluded from our conclusions (see Section “Discussion”).

Bottom Line: For diploid taxa, a Bayesian concordance analysis and a coalescent-based dated species tree was inferred from all gene trees.The resulting multilocus phylogeny reveals for the first time species phylogeny and progenitor-derivative relationships of all di- and polyploid Hordeum taxa within a single analysis.Our study proves that it is possible to obtain a multilocus species-level phylogeny for di- and polyploid taxa by combining PCR with next-generation sequencing, without cloning and without creating a heavy load of sequence data.

View Article: PubMed Central - PubMed

Affiliation: Institute of Plant Genetics and Crop Plant Research (IPK), D-06466 Gatersleben, Germany; brassac@ipk-gatersleben.de.

Show MeSH