Reticulate evolution in Ranunculus cantonensis polyploid complex and its allied species

Hybridization is particularly likely to occur in initial and young polyploid complexes, and interspecies hybridization between diverged species usually leads to a complicated reticulate evolution. The Ranunculus cantoniensis complex and its allied species include R. chinensis (2x), R. silerifolius var. silerifolius (2x), R. cantoniensis (4x), R. trigonus (2x), R. shuichengensis (2x), R. diffusus (4x), R. repens (4x), R. vaginatus (5x) and R. sieboldii (6x, 8x). Many morphological intermediates can be found between the members of this complex, and the relationship among members is complicated. By analyzing internal transcribed spacers and nrDNA FISH (fluorescence in situ hybridization) signals, we unraveled the phylogenetic and genetic constitution of the various taxonomic units of this complex. Haplotypes were highly separated by median-joining network analysis and at least four haplogroups emerged in which there were 11 primary haplotypes; six out of ten taxa shared haplotype 1, suggesting that haplotype 1, a variation of the primary haplotype R. chinensis, served as the pivotal genome in the complex. The pollen characteristics and electrophoretic patterns of R. vaginatus (5x) showed it to be an intermediate between R. diffusus (4x) and R. sieboldii (6x). The distribution of R. vaginatus (5x) was located at the junction of the distributions of R. diffusus (4x) and R. sieboldii (6x). Ranunculus vaginatus (5x) shared haplotypes 7 and 8 with R. diffusus (4x), and haplotypes 8 and 9 with R. sieboldii (6x). This proved that R. vaginatus (5x) emerged from hybridization between R. diffusus (4x) and R. sieboldii (6x). The results of FISH also support a hybrid origin of R. vaginatus (5x). The findings of this study clearly show that there are only eight taxa in this polyploid complex including R. chinensis (2x), R. silerifolius var. silerifolius (2x), R. trigonus (2x), R. silerifolius var. dolicanthus(2x), R. cantoniensis (4x), R. diffusus (2x), R. vaginatus (5x) and R. sieboldii (6x, 8x). These taxa associated with each other by hybridizing with the pivotal genome. Ranunculus cantoniensis (4x) and R. vaginatus (5x) arose from hybridization events between diverged species in the polyploid complex, leading to a complicated reticulate evolution.