Maximum likelihood analysis of the protein coding genes.

<p>The maximum likelihood analysis of the mitochondrial protein coding genes of six annelids shows that branch lengths among them are similar, suggesting that <i>Nephtys</i> does not have an obviously slower rate that might create a propensity for harboring introns.</p

Primers used for completion of <i>Nephtys</i>' mtDNA amplification.

<p>Primers used for completion of <i>Nephtys</i>' mtDNA amplification.</p

Key characteristics of group I and group II self-splicing introns.

<p>Key characteristics of group I and group II self-splicing introns.</p

Phylogenetic analysis of 71 group II intron ORFs. A maximum likelihood analysis of the amino acid sequence for 71 ORFs suggests the <i>cox1</i> ORF718 of the marine centric diatom <i>Thalassiosira pseudonana</i> as sister to the <i>Nephtys</i>'s ORF.

<p>Red stars indicate a bootstrap support ≥90. Names of taxa are indicated by the capital letter of the genus name, followed by species name and when applicable the intron location (specified in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0001488#pone-0001488-t003" target="_blank">table 3</a>).</p

Mitochondrial, chloroplast and bacterial group II introns included in the phylogenetic analysis (modified from Zimmerly et al. ).

<p>Mitochondrial, chloroplast and bacterial group II introns included in the phylogenetic analysis (modified from Zimmerly et al. ).</p

Results of Branch-sites analyses.

<p>Foreground (<i>Cuscuta nitida</i> or <i>Epifagus virginiana</i> lineages) and background omega (ω = <i>dN/dS</i>) parameters for Branch-sites models with significant LRT results (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0005982#pone-0005982-t002" target="_blank">Table 2</a>).</p>*<p>Background and foreground values of ω₀, ω₁ and ω₂ listed in the table.</p>**<p>Sites implicated as evolving under positive selection in Positive Selection bXs model (Bayes empirical Bayes posterior probabilities >0.95 <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0005982#pone.0005982-Yang2" target="_blank">[25]</a> in tests of positive selection on foreground branches (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0005982#pone-0005982-t002" target="_blank">Table 2</a>) listed as position in alignment and derived amino acid residue.</p

Phylogenies of Convolvulaceae (a) and Orobanchaceae (b) inferred from full and partial nucleotide sequences of <i>matK</i>.

<p>Branches shown in red denote significant increases in selection within domain X of <i>matK</i>. Trees were produced using Maximum Likelihood with bootstrap values (100 replications) shown at the nodes (bootstrap values of 100 are denoted by asterisks). Taxonomic delimitations of <i>Cuscuta</i> subgenera and Convolvulaceae are boxed and labeled. Group IIA intron losses are mapped on branches where they are inferred to have occurred. The branch joining <i>Ipomoea</i> with <i>Cuscuta</i> species, denoted with a dotted line, was collapsed for analysis due to low support in other studies.</p

Intron distribution in relevant taxa.

*<p>trnV introns in <i>Cuscuta</i> subg. <i>Monogyna</i> have deletions that may render them pseudogenes.</p><p>Intron presence or absence is shown for <i>Nicotiana tabacum</i> (Solanaceae), <i>Ipomoea purpurea</i> and <i>Cuscuta</i> spp. (Convolvulaceae), and <i>Epifagus virginiana</i> (Orobanchaceae). <i>Nicotiana</i>, <i>Ipomoea</i>, and <i>Cuscuta</i> are classified in the order Solanales, while <i>Epifagus virginiana</i> is in the closely related order Lamiales. Subgeneric taxonomic classifications are listed for <i>Cuscuta</i> spp. <i>Nicotiana</i> intron distribution is typical of most angiosperms. “+” indicates intron present, “−” indicates precise intron loss from an intact gene, and “X” indicates loss of gene (and intron) from the plastid genome. Intron data for <i>Nicotiana</i>, <i>Ipomoea</i>, <i>Cuscuta exaltata</i>, <i>Cuscuta reflexa</i>, <i>Cuscuta gronovii</i>, <i>Cuscuta obtusiflora</i>, and <i>Epifagus</i> were gleaned from complete genome sequences available on genbank; all other data are based on PCR and PCR sequencing assays.</p

Primer sequences designed for this study.

<p><i>matK</i> primers designed using sequence from subgenus <i>Monogyna</i> are designated by the suffix subgM, ones designed using subgenus <i>Cuscuta</i> sequences by subgC, and ones designed with Convolvulaceae sequences by Conv.</p

Voucher information and <i>matK</i> GenBank accession numbers.

<p>Sequences generated outside of our group are shown in bold. Specimens that lacked enough material for herbarium voucher are denoted by an asterisk (*); photographs of the dissected flowers used for identification are available upon request. Plant material or DNA from other labs where no voucher information was provided were verified by sequence identity to existing vouchered sequences on GenBank and are marked with a #.</p