DPMT validation for MOB_P3 and MOB_P4 relaxases.

<p>A) Phylogenetic tree of MOB_P3 and MOB_P4 relaxase families. B) Alignment of the relaxase motifs used to design the MOB_P3 and MOB_P4 degenerate primers (P3-f+P3-r, continuous black; and P4-f+P4-r, continuous dark grey). C) Amplicons obtained with primers for subfamily MOB_P31 (P3-f and P3-r). D) Amplicons obtained with primers for subfamily MOB_P42 (P4-f and P4-r). Symbols, colour codes and lanes as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0040438#pone-0040438-g001" target="_blank">Figure 1</a>.</p

DPMT validation for MOB_Q relaxases.

<p>A) Phylogenetic tree of MOB_Q relaxase family. B) Alignment of the relaxase motifs used to design the MOB_Q degenerate primers (Q11-f+Q11-r, continuous black; Q12-f+Q12-r, continuous dark grey; and Qu-f+Qu-r, continuous grey). C) Amplicons obtained with primers for subfamily MOB_Q11 (Q11-f and Q11-r). D) Amplicons obtained with primers for subfamily MOB_Q12 (Q12-f and Q12-r). E) Amplicons obtained with primers for subfamily MOB_Qu (Qu-f and Qu-r). Symbols, colour codes and lanes as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0040438#pone-0040438-g001" target="_blank">Figure 1</a>.</p

DPMT validation for MOB_C relaxases.

<p>A) Phylogenetic tree of MOB_C relaxase family. B) Alignment of the relaxase motifs used to design the MOB_C degenerate primers (C11-f+C11-r, continuous black; C12-f+C12-r, continuous dark grey). C) Amplicons obtained with primers for subfamily MOB_C11 (C11-f and C11-r). D) Amplicons obtained with primers for subfamily MOB_C12 (C12-f and C12-r). Symbols, colour codes and lanes as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0040438#pone-0040438-g001" target="_blank">Figure 1</a>.</p

DPMT validation for MOB_P1 relaxases.

<p>A) Phylogenetic tree of MOB_P1 relaxases. B) Alignment of the relaxase motifs used to design the MOB_P1 degenerate primers (P11-f, continuous black; P12-f, continuous dark grey; P131-f, continuous grey; P14-f, continuous light grey; and P1-r, dashed black). C) Amplicons obtained with primers for subfamily MOB_P11 (P11-f and P1-r). D) Amplicons obtained with primers for subfamily MOB_P12 (P12-f and P1-r). E) Amplicons obtained with primers for subfamily MOB_P13 (P131-f and P1-r). F) Amplicons obtained with primers for subfamily MOB_P14 (P14-f and P1-r). Symbols, colour codes and lanes as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0040438#pone-0040438-g001" target="_blank">Figure 1</a>.</p

List of degenerate primers used for DPMT.

a<p>For each oligonucleotide, CORE nucleotides are in bold and CLAMP sequences in normal lettering. Underlined codons do not encompass all the possible variability to avoid excessive degeneracy. The sequences used are biased to accommodate the DNA sequences of existing elements.</p>b<p>Prototype plasmid for the given MOB subfamily.</p>c<p>Amplicon size obtained from the prototype plasmid relaxase gene.</p>d<p>Nucleotide coordinates of the prototype plasmid contained in the corresponding amplicon. An asterisk indicates that the relaxase gene is encoded in the complementary strand.</p

DPMT validation for MOB_P5 relaxases.

<p>A) Phylogenetic tree of MOB_P5 relaxase family. B) Alignment of the relaxase motifs used to design the MOB_P5 degenerate primers (P51-f, continuous black; P52-f, continuous dark grey; P5-r, dashed black; and P53-f+P53-r, continuous grey) C) Amplicons obtained with primers for subfamily MOB_P51 (P51-f and P5-r). D) Amplicons obtained with primers for subfamily MOB_P52 (P52-f and P5-r). E) Amplicons obtained with primers for subfamily MOB_P53 (P53-f and P53-r). Symbols, colour codes and lanes as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0040438#pone-0040438-g001" target="_blank">Figure 1</a>.</p

Relaxases found in two test collections.

a<p>Isolate collections analyzed with DPMT.</p

DPMT validation for MOB_H relaxases.

<p>A) Phylogenetic tree of MOB_H relaxase family. B) Alignment of the relaxase motifs used to design the MOB_H degenerate primers (H11-f+H11-r, continuous black; H121-f+H121-r, continuous dark grey; and H2-f+H2-r, continuous grey). C) Amplicons obtained with primers for subfamily MOB_H11 (H11-f and H11-r). D) Amplicons obtained with primers for subfamily MOB_H121 (H121-f and H121-r). E) Amplicons obtained with primers for subfamily MOB_H2 (H2-f and H2-r). Symbols, colour codes and lanes as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0040438#pone-0040438-g001" target="_blank">Figure 1</a>.</p

Plasmids and relaxase genes used as controls in validation experiments.

a<p>Accession number of the transmissible plasmid encoding the corresponding relaxase gene.</p>b<p>Nucleotide coordinates of the cloned relaxase fragment in the accession number of the original plasmid. An asterisk indicates that the relaxase gene is coded in the complementary strand of the original plasmid sequence.</p>c<p>Incompatibility group of the wt plasmid.</p>d<p>MOB subfamily of each relaxase gene.</p

Correspondence between MOB and Rep types.

<p>A) Simplified phylogenetic representation of the five relaxase MOB families considered in this study. Coloured triangles represent the MOB subfamilies amplified by DPTM. Their width and depth correspond, respectively, to the abundance and phylogenetic diversity of their relaxase sequences (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0040438#pone.0040438.s001" target="_blank">Table S1</a>). B) The Inc groups contained within each MOB subfamily are indicated at the right, boxed in the same colour. When no Inc group is contained, the name of a prototype plasmid is given.</p