Cumulative distribution plots of the between-groups alignment scores for 50 of the 51 chloroplast proteins of monocots and eudicots (the plot for psbF is Fig 2A).

<p>The ancestral alignment score (s(<i>a_X</i>,<i>a_Y</i>)) is indicated by a small circle on each plot. There are 1056 comparisons (24 monocots×44 eudicots) for each protein. The y-axis is the same for each gene, but the x-axis is strongly dependent on the length of the protein (see also <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0069924#pone-0069924-g004" target="_blank">Figure 4</a>).</p

Results for the 51 genes for the monocot/eudicot chloroplast dataset, and with the ancestral sequences (a_x and a_y) inferred independently.

<p>Results for the 51 genes for the monocot/eudicot chloroplast dataset, and with the ancestral sequences (a_x and a_y) inferred independently.</p

We use two natural subgroups (X and Y), independently align the sequences for the species in each subgroup, independently determine the optimal tree for each subgroup, independently infer the ancestral sequences a_x and a_y on the optimal subtrees (in practice the sequence at the nearest node to the root of the subtree is estimated), and finally measure the pairwise alignment score between the ancestral sequences, s(a_x,a_y).

<p>Separately, we measure the alignment score between each pair of sequences (s(_i,j)) with one member in each of the two subsets, for example, s(_a,k), s(_a,l), s(_a,m), and so on.</p

Cumulative frequency plots comparing the alignment score for the ancestral sequences (s(a_x,a_y), small circle) with the alignment scores of all pairs of proteins, s(<i>i</i>,<i>j</i>).

<p>The example is the monocot/eudicot chloroplast dataset and for the short protein psbF (<b>2A</b>), a longer protein atpA (<b>2B</b>), and the 51 concatenated genes (<b>2C</b>). The x-axis shows the alignment score, which increases with the length of the protein(s), and is largest for the 51 concatenated proteins. There are 1056 s(<i>i</i>,<i>j</i>) scores between pairs of 24 monocots and 44 eudicots, and the y-axis indicates where the s(a_x,a_y) fits as a proportion of this number. For some short proteins in particular, multiple s(<i>i</i>,<i>j</i>) values equal the ancestral score s(a_x,a_y), and in this case our test conservatively places the ancestral score below the rest (as in psbF in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0069924#pone-0069924-g002" target="_blank">Fig 2A</a>).</p

Protein length versus proportion of pairwise alignment scores higher than the ancestral score, for 4 datasets.

<p>Because of the possibility of slightly different gene lengths just one of the two datasets is used for illustration. As expected, longer proteins show convergence more strongly. Chloroplast results (4A and 4B) are for 51 chloroplast genes for divergences of monocots and for angiosperms (flowering plants). There are 7 nuclear proteins for 4C and 12 for the mitochondrial data in 4D.</p

Calculation of alignment score for the inferred ancestral monocot and eudicot sequences of the psbK gene.

<p>The ancestral sequences for the monocots, and the eudicots, are inferred independently, and then the alignment scores calculated in the program MUSCLE. The individual column scores depend on the frequencies and properties of the two amino acids; higher scores are given for pairs that are similar (readily substitutable) or specific (more readily substitutable for each other than for other amino acids). The column scores are summed to produce the alignment score.</p

Additional file 1: of The Antarctic sea ice alga Chlamydomonas sp. ICE-L provides insights into adaptive patterns of chloroplast evolution

Table S1. Results of the basic (M0) model test of individual genes and the concatenated data. Table S2. Results of the correct P-value of LRT in branch-model test. Table S3. Results of site-model test to each gene-specific and the concatenated data. Table S4. Results of branch-site model analyses using the concatenated data. Table S5. Results of convergence test between Chlamydomonas sp. ICE-L and Dunaliella salina. Figure S1. Results of divergence time analyses. The estimated divergence times were showed with 95% confidence intervals. Figure S2. The dN/dS estimates for the concatenated data using the M8 random-site model. The y-axis shows the BEB posterior mean estimate of dN/dS for each site. (DOCX 341 kb

S2 - S2.txt

alignment of first and the second codon positions 25246 positions in lengt

S6 - S6.zip

zip archive containing scripts used in the pape

S5 - S5.pdf

NNet split networks for S1 and S4 alignment