Figure 1.
Phylogenetic Distribution of Hairpin Numbers for ITS Secondary Structures.
A tribal level phylogeny of the Brassicaceae, strict consensus tree of the 200 most parsimonious trees estimated with ITS sequences [2], was utilized to investigate the evolution of the number of hairpins present in the secondary structures of both ITS1 and ITS2. Bootstrap support values greater than 60% are shown above branches [2]. It is notable that the ITS tree does neither fully reflect the tribal phylogeny nor is at any deep node highly significantly supported, but overall-topology is in congruence with multi-locus phylogenies considering major lineages [7], [8]. Tribes not assigned to one of the three major lineages are actually combined with an "expanded lineage II" [46], which might have to be revised in future. The three major phylogenetic lineages are shown within colored blocks with Lineage I (orange), Lineage II (blue) and Lineage III (green). The number of hairpins for each secondary structure is shown at the phylogenetic tips with 3 (orange boxes), 4 (yellow boxes), 5 (green boxes), 6 (blue boxes), and 7 (purple boxes). Tribes with a lack of available complete ITS1 data are marked as 'NA'. Tribes with secondary structures with different number of total hairpins from different species are also indicated (e.g. Camelineae 6/7 for ITS1; 6 and 7 hairpin structures are observed) within the colored box of the fewest hairpined structure. Examples of secondary structures are shown (top-bottom order): 1. Anchonium billardierei ITS1 (Anchonieae), 2. Aethionema arabicum ITS1 (Aethionemeae), 3. Halimolobos lasiolaba ITS2 (Halimolobeae), and 4. Arabis scabra (Arabideae).
Figure 2.
Structure of the rDNA region in Plants.
An illustration of the nucleolus organizing region (NOR), shown as red colored region on chromosome, is associated with forming the nucleolus and site for the biosynthesis of the components of the ribosome. The NOR region contains hundreds of tandem duplicated copies of rDNA gene clusters (depicted as yellow rectangles), and each gene cluster consists of seven main components including two internal transcribed spacers (i.e. ITS1 and ITS2). These ITS regions form self-splicing secondary structures as transcribed products. Shown is the 5 hairpin structure for ITS1 and the 3 hairpin structure for ITS2.
Figure 3.
Hairpin Size Distributions for ITS1 and ITS2 Secondary Structures.
Panel A shows the frequency of the total percentage of paired bases for all ITS1 (Blue) and ITS2 (Red) sequences. The majority of both ITS regions have over 50% of positions paired with other sequence positions to form secondary structures. Panel B shows the frequency of the total number of hairpins for all the ITS1 (Blue) and ITS2 (Red) structures. These data are for the lowest energy state structure for each ITS sequence (see Supplemental File 1).
Figure 4.
Comparison of Secondary Structures of 100 ITS and 100 Random Sequences.
A scatter plot of the lowest energy state values (x-axis) and all possible secondary structures (y-axis) for 50 ITS1 (Blue Diamonds), 50 ITS2 (Red Squares) and 100 randomly generated sequences (Green Triangles) (Supplemental File 1) estimated using RNAstructure 5.3 (Reuter and Mathews, 2010).