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Potamogeton  is one of the most difficult groups to clarify in aquatic plants, which has an extensive range of interspecific morphological and ecological diversity. Internal transcribed spacer (ITS) is prevalent for phylogenetic analysis in plants. However, most researches demonstrate that ITS has a high percentage of homoplasy in phylogenetic datasets. In this study, eighteen materials were collected in  Potamogeton  from China and incongruence was shown between the  rbcL  and ITS phylogenies. To solve the discrepancy, we employed a novel barcode  PHYB  to improve resolution and accuracy of the phylogenetic relationships. The  PHYB  phylogeny successfully resolved the incongruence between the  rbcL  and ITS phylogenies. In addition, six hybrids were confirmed using  PHYB , including  P .  compressus  ×  P .  pusillus ,  P .  octandrus  ×  P .  oxyphyllus ,  P .  gramineus  ×  P .  lucens ,  P .  distinctus  ×  P .  natans ,  P .  distinctus  ×  P .  wrightii , and  S .  pectinata  ×  S .  amblyophylla . Whereas, only one hybrid was identified ( P .  compressus  ×  P .  pusillus ) by ITS, indicating that ITS homoplasy was present in  Potamogeton  and ITS was completely homogenized to one parental lineage. Thus, ITS might have limited utility for phylogenetic relationships in  Potamogeton . It is recommended that a three-locus combination of chloroplast DNA gene, ITS and  PHYB  is potential to effectively reveal more robust phylogenetic relationships and species identification.

Identification of Hybrids in Potamogeton: Incongruence between Plastid and ITS Regions Solved by a Novel Barcoding Marker PHYB