EmsB PCR amplification was performed as previously reported (17 (link)). Capillary electrophoresis of PCR products was performed on a 3500 genetic analyzer (Life Technologies, Foster City, CA, USA). The size and height of each peak of the electrophoretic profile constituting the EmsB profiles were determined with the use of GeneMapper 4.1. The characterization of EmsB profiles composed of several peaks or alleles from 209 to 241 bp was carried out as previously described (11 (link), 20 (link)). The hierarchical clustering analysis was done using the Euclidean distance and the average link clustering method (UPGMA) (22 ). The uncertainty of clusters was evaluated by multiscale bootstrap resampling (B = 1,000) and given by approximately unbiased p-values (AU), according to Shimodaira (23 (link), 24 ). Clustering analyses were performed using R statistical software (25 ) and the pvclust library (26 (link)). In each dendrogram, EmsB microsatellite data from previously genotyped samples from the Arctic and Asian groups (11 (link), 13 (link)) were added. The genetic threshold of 0.08 was used to determine the genotyping status of each sample (11 (link)), while two E. granulosus sensu stricto (G1) samples were used as the outgroup.
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