Methylation sensitive AFLP data of Dactylorhiza allopolyploids

The data is provided in four tabs: the "Original" tab contains the initial information obtained from the two paired profiles per individual (i.e., MspI and HpaII); the "Unmethylated" tab includes fragments present in both profiles simultaneously; the "MeCpG" tab includes the information from MspI; the "hemiMeCpCpG" tab includes the information from HpaII. Redundant information between the last 3 tabs has been retained only once

AFLP data for Ranunculus alpestris group

AFLP data for Ranunculus alpestris grou

R_script_Mantel_RMA

R script for running a Mantel test and a Reduced Major Axis (RMA) regression

ITS Polymorphisms Shed Light on Hybrid Evolution in Apomictic Plants: A Case Study on the <i>Ranunculus auricomus</i> Complex

<div><p>The reconstruction of reticulate evolutionary histories in plants is still a major methodological challenge. Sequences of the ITS nrDNA are a popular marker to analyze hybrid relationships, but variation of this multicopy spacer region is affected by concerted evolution, high intraindividual polymorphism, and shifts in mode of reproduction. The relevance of changes in secondary structure is still under dispute. We aim to shed light on the extent of polymorphism within and between sexual species and their putative natural as well as synthetic hybrid derivatives in the <i>Ranunculus auricomus</i> complex to test morphology-based hypotheses of hybrid origin and parentage of taxa. We employed direct sequencing of ITS nrDNA from 68 individuals representing three sexuals, their synthetic hybrids and one sympatric natural apomict, as well as cloning of ITS copies in four representative individuals, RNA secondary structure analysis, and landmark geometric morphometric analysis on leaves. Phylogenetic network analyses indicate additivity of parental ITS variants in both synthetic and natural hybrids. The triploid synthetic hybrids are genetically much closer to their maternal progenitors, probably due to ploidy dosage effects, although exhibiting a paternal-like leaf morphology. The natural hybrids are genetically and morphologically closer to the putative paternal progenitor species. Secondary structures of ITS1-5.8S-ITS2 were rather conserved in all taxa. The observed similarities in ITS polymorphisms suggest that the natural apomict <i>R. variabilis</i> is an ancient hybrid of the diploid sexual species <i>R. notabilis</i> and the sexual species <i>R. cassubicifolius</i>. The additivity pattern shared by <i>R. variabilis</i> and the synthetic hybrids supports an evolutionary and biogeographical scenario that <i>R. variabilis</i> originated from ancient hybridization. Concerted evolution of ITS copies in <i>R. variabilis</i> is incomplete, probably due to a shift to asexual reproduction. Under the condition of comprehensive inter- and intraspecific sampling, ITS polymorphisms are powerful for elucidating reticulate evolutionary histories.</p></div

Data

Source data used. The separate worksheets refer to information related to: (1) microsatellite data, (2) pairwise geographical and genetic distances of D. majalis, (3) pairwise geographical and genetic distances of D. traunsteineri, (4) population pairwise GST data and (5) additional parameters for analysis

R_script_pwGST

R script to calculate pairwise Gst analyses

List of directly sequenced individuals.

a)<p><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0103003#pone.0103003-Hrandl14" target="_blank">[96]</a>.</p>b)<p><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0103003#pone.0103003-Hrandl2" target="_blank">[46]</a>.</p>c)<p>functional sexual seed, but with aposporous initials (for details see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0103003#pone.0103003-Hojsgaard1" target="_blank">[58]</a>).</p>d)<p>functional sexual seed, but with low rates of aposporous seeds (for details see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0103003#pone.0103003-Hojsgaard1" target="_blank">[58]</a>).</p

Geometric morphometric analysis of individuals exhibiting the characteristic “<i>auricomus</i>” morphology.

<p>(A) An example of the sampled fresh plant material for DNA sequencing and geometric morphometric analyses. (B) 2D landmarks digitalization on the leaf outline. (C) Principal components analysis of the shape variables (i.e., Relative warps analysis, RWA) extracted from <i>R. notabilis</i> (blue squares), <i>R. cassubicifolius</i> × <i>notabilis</i> (red crosses) and <i>R. variabilis</i> (violet triangles) 2D landmark data. (D) Mean shapes of <i>R. notabilis</i>, <i>R. cassubicifolius</i> and <i>R. variabilis</i> reconstructed from each centroid (visualized as symbols surrounded by black-lined circles) of the three scatter clusters shown in (C). The between-group differences were tested by permutation tests (lower left triangle, ***  =  p<0.001) and the distances between the mean shapes are expressed as Procrustes distances (upper right triangle).</p

NeighborNet analysis of interspecific ITS1+ITS2 variability within the <i>Ranunculus auricomus</i> complex.

<p>NeighborNet analysis of all ITS1+ITS2 sequences obtained by direct sequencing of the studied individuals. The spring leaf silhouettes illustrate the main phenotypic differences between the two morphotypes: the <i>auricomus</i>-morphotype (characteristic for <i>R. notabilis</i>, <i>R. variabilis</i> and the synthetic hybrid <i>R. cassubicifolius</i> × <i>notabilis</i>) and the <i>cassubicus</i>–morphotype (i.e., <i>R. carpaticola</i> and <i>R. cassubicifolius</i>). Individuals belonging to <i>R. carpaticola</i> are marked as “P”, <i>R. cassubicifolius</i> as “S”, <i>R. notabilis</i> as “N” and <i>R. variabilis</i> as “V”, respectively. Identical sequences representing the same ribotype are listed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0103003#pone-0103003-g003" target="_blank">Figure 3</a>. Bootstrap values are given for the main clusters.</p

5.8S secondary structure model for <i>Ranunculus notabilis</i> 5613-1.

<p>(A) The partial 5.8S secondary structure model comprising conserved sequence regions (M1-M3), conserved helices (B5-B8) and highlighted sites affected by nucleotide substitutions (for the whole 5.8S secondary structure see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0103003#pone.0103003.s001" target="_blank">Figure S1</a>). (B) A summary Table of 5.8S nucleotide polymorphisms detected in clones.</p