Abstract:

In order to revealed the genetic resources and Germplasm repository of the clam Paphia undulate. The present studies reported that isolation and screening of microsatellite markers from the clam Paphia undulate were performed using the method of Fast Isolation by AFLP of Sequences Containing Repeats (FIASCO). After restriction digests of genomic DNA with Mse I, the digested fragments were ligated with adaptors and then hybridized with 5’-biotinylated (CA)15 probe or 5’-biotinylated (AAG)7 probe. The tentative microsatellite DNA was isolated by streptavidin-coated magnetic beads from the hybridized mixture. After purification, the isolated microsatellite DNA was amplified using degenerated primer Mse P and then cloned into T-vector. After transforming, the microsatellite- enriched library was constructed. The second PCR screening was performed using the primer of (CA)15 probe or (AAG)7 probe) and T vector’s primers. The SSR developed were used for analyzing genetic diversity of Zhanjiang wild population of the species. A total of 108 alleles were detected by the 8 SSR loci. The number of alleles per SSR locus ranged from 5 to 19 in a sample of 39 individuals captured in inshore water of Zhanjiang. Observed and expected heterozygosity per locus varied from 0.400 to 0.882 and from 0.666 to 0.926, respectively. Four loci (Pun4, Pun5, Pun6，Pun7) showed significant departure from Hardy–Weinberg equilibrium after sequential Bonferroni correction (P<0.00625). All loci were high polymorphic loci. This study demonstrated that FIASCO is a useful technique of microsatellite isolation for the species. 8 microsatellite markers should provide a tool for assessing genetic diversity and population structure of wild or breeding stocks.