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Genetic variation of nuclear ribosomal ITS (nrITS) and chloroplast DNA (cpDNA) regions was investigated in Brasenia schreberi (Cabombaceae) to assess the population structure and to infer the evolutionary relationship among 12 populations distributed in South Korea. The combined sequence of the two regions was aligned to 2,069 bp, of which 28 sites were variable. In total, 20 genotypes were identified from 240 individuals of B. schreberi. Genotype diversity (Gd) and nucleotide diversity (Pi) on Jeju Island (Gd = 0.2511, Pi = 0.00012) were higher than those of the mainland of South Korea (Gd = 0.1358, Pi = 0.00005). The relatively low level of genetic variation of the mainland populations is associated with its higher genetic differentiation (G ST = 0.095 on mainland and 0.039 on Jeju Island) and human activities. Minimum spanning network analysis demonstrated that the investigated populations of B. schreberi were subdivided into two geographical groups: the mainland of South Korea and Jeju Island. In addition, analysis of molecular variation showed that a large proportion (73.55%) of genetic differentiation existed between the two regions. These results strongly suggest the presence of significant barriers to gene flow between regions. Thus, the management unit for B. schreberi should be carefully designed to avoid the potential risk that can results from the admixture of individuals from the mainland and Jeju Island regions.

On the basis of amplified fragment length polymorphism (AFLP) and nucleotide sequence data from nuclear ribosomal internal transcribed spacer (nrITS) and three chloroplast DNA regions (rbcL, cpITS, and trnS-psbC spacer), we investigated the species delimitation and the evolutionary lineage of Isoetes asiatica from Hokkaido, Japan. The neighbor-joining (NJ) dendrogram based on AFLP markers revealed the well-defined clusters (bootstrap value = 100%) of I. asiatica. Results from the principal component analysis are largely congruent with those obtained in the NJ dendrogram. The maximum parsimony analysis, based on data from nrITS and three chloroplast DNA sequences, supported a monophyly of three species, I. asiatica, Isoetes echinospora, and Isoetes maritima from Hokkaido, Kamchatka, and Alaska regions. The distinct species status of I. asiatica was also well supported in the combined chloroplast DNA phylogeny. Therefore, I. asiatica appear to represent example of gradual speciation due to spatial isolation of ancestral populations followed by genetic divergence. Our results also suggest that I. asiatica is probably not the ancestral diploid of the polyploids occurring in East Asia.

Brasenia schreberi J.F. Gmelin is a declared endangered species found in the lakes and ponds of South Korea. For planning its conservation strategy, we examined the genetic diversity within and among six populations, using randomly amplified polymorphic DNA (RAPD) and amplified fragment length polymorphism (AFLP). Polymorphisms were more frequently detected per loci with AFLP (69.3%) than RAPD (36.8%). High genetic diversity was recognized within populations: polymorphic loci (PPL) values ranged from 36.3% in the CJM population to 74.5% in the GGT population, with a mean value of 47.8% based on AFLP markers. Great genetic differentiation (θB) was detected among the six populations (0.670 on RAPD and 0.196 on AFLP), and we calculated a low rate of gene flow (Nem), i.e., 0.116 on RAPD and 0.977 on AFLP. Furthermore, a Mantel test revealed that no correlation existed between genetic distances and geographical distances among the six local populations, based on RAPD or AFLP markers. These results are attributed to a number of factors, including an insufficient length of time for genetic diversity to be reduced following a natural decline in population size and isolation, adaptation of the genetic system to small population conditions, and a restricted gene flow rate. Based on both its genetic diversity and population structure, we suggest that a strategy for conserving and restoringB. schreberi must focus on maintaining historical processes, such as high levels of outbreeding, while monitoring increased gene flow among populations. This is because a reduction in genetic diversity as a result of genetic drift is undesirable.