SSR Isolation And Population Genetic Analysis Of PAA Spinosa

Many populations of amphibians throughout the world have declined markedly, some species have become extinct. The conservation of amphibians is faced severe situation. Amphibians are suitable for the researches on population dynamics and conservation biology and always are used as indicators of ecological environmental monitoring. In china, determination and evaluation of biodiversity protection priority area for many frogs are both lack of fundamental data. The endemic species of China can provide specific informations of amphibian population ecology. The giant spiny frog (Paa spinosa), distributed in the south of China and north of Vietnam, was a threatened species with commercial and nutritious importance in China and listed on IUCN Red List Categories as "Vulnerable" (A2abc). Genetic markers and population genetics study are helpful to the protection of P. spinosa, they are also the basic of Chinese amphibian Phylogeography. The 13 geographical populations of the frog were select, their genetic diversity and structure were assessed with the microsatellite markers. The results are as follows:Fifteen polymorphic microsatellite loci for the giant spiny frog P. spinosa, were characterized. These loci were highly polymorphic when screened in 38 individuals from two different populations, with 9-23 alleles per locus. The range of observed and expected heterozygosity was 0.393-0.906 and 0.432-0.955, respectively. These polymorphic loci will be useful for assessing genetic diversity, population structure, gene flow, population assignment, and determining paternity in the giant spiny frog.We investigated genetic diversity and population genetic structure by using 12 polymorphic microsatellite loci based on 210 samples from 13 populations from its entire range. A total of 206 microsatellite alleles were observed, and the mean expected heterozygosity (H_E) within each locality was 0.78 (range: 0.676-0.892). Genetic differentiation was high (overall F_ST = 0.114) and analyses based on a Bayesian clustering method revealed that the dataset constituted three genetic clusters. Also, the neighbour-joining analysis yielded a phylogenetic tree with high bootstrap values on three internal branches, defining three distinct genetic clusters corresponding to Western China, Central China and Eastern China. Analysis of molecular variance (AMOVA) consistently supported that these clusters have highly differentiated without gene flow with one another and fixed differences were observed at all locus. Indications of a genetic bottleneck were found in five populations. The microsatellite data clearly distinguished these three genetic clusters. Although confirmation of the status of three genetic clusters in the P. spinosa as separate species requires further data, our study supports the definition of these clusters as different evolutionary significant units.