Abstract:In order to evaluate the genetic diversity of sea cucumber Apostichopus japonicus and breed eugenic varieties, a CA microsatellite containing short fragment genomic DNA library of sea cucumber was constructed using the FIASCO(Fast Isolation by AFLP Sequences Containing Repeats)method. In 140 randomly selected and sequenced clones, 118 contained a CA repeat motif. According to Weber(1990) classification rules, the sequences were divided into three categories: 83 perfect repeat sequences without interruptions in the runs of CA or GT dinucleotides (70.4% of total), 30 imperfect repeat sequences with one or more interruptions in the run of repeats (25.4%), and 5 compound repeat sequences with adjacent tandem simple repeats of a different sequence (4.2%). Twenty pairs of primers were designed and used to investigate the polymorphism and genetic structure of wild A. japonicus collected from China(CH), Korea(KW, KE), Japan(JA) and Russia(RU). Cross amplification of twenty loci were also tested in Parastichopus parvimensis which were collected in USA(AM). The results demonstrated that all the markers were polymorphic and 16 loci had high polymorphism (PIC>0.5). The average observed and expected heterozygosities of the 20 loci were 0.39 and 0.69, respectively. In addition, a total of 231 alleles (102 effective alleles) were detected at 20 loci in the total samples and the alleles number of each locus ranges from 3 to 20 with average of 11.6. The Hardy-Weinberg Equilibrium (HWE) analysis showed that all the loci deviated from genetic equilibrium in the six populations and heterozygote deficiency occurred to different extent in total samples (Fis>0). Genetic identity of the five A. japonicus populations was 0.71, while the genetic identity between A. japonicus and P. parvimensis were quiet low. In the UPMGA tree, CH and KW populations formed a cluster, RU, JA and KE populations formed another cluster, while AM felt into a seperate cluster. The result of cluster analysis indicated that the clustering order had significant correlation with geographic distance.
