ITS non-concerted evolution and rampant hybridization in the legume genus Lespedeza (Fabaceae)

The internal transcribed spacer (ITS) as one part of nuclear ribosomal DNA is one of the most extensively sequenced molecular markers in plant systematics. The ITS repeats generally exhibit high-level within-individual homogeneity, while relatively small-scale polymorphism of ITS copies within individuals has often been reported in literature. Here, we identified large-scale polymorphism of ITS copies within individuals in the legume genus Lespedeza (Fabaceae). Divergent paralogs of ITS sequences, including putative pseudogenes, recombinants, and multiple functional ITS copies were sometimes detected in the same individual. Thirty-seven ITS pseudogenes could be easily detected according to nucleotide changes in conserved 5.8S motives, the significantly lower GC contents in at least one of three regions, and the lost ability of 5.8S rDNA sequence to fold into a conserved secondary structure. The distribution patterns of the putative functional clones were highly different between the traditionally recognized two subgenera, suggesting different rates of concerted evolution in two subgenera which could be attributable to their different extents/frequencies of hybridization, confirmed by our analysis of the single-copy nuclear gene PGK. These findings have significant implications in using ITS marker for reconstructing phylogeny and studying hybridization.