Isolation And Characterization Of Polymorphic Microsatellite Loci In Olive Flounder (Paralichthys Olivaceus)

Olive flounder (Paralichthys olivaceus) is a commercially important aquaculture fish species that is being widely cultured in China. The isolation and using of microsatellite in Olive flounder are vacant in China, which is not fit for the development of the aquaculture of Oliver flounder. In this study we using two methods to obtain the microsatellite markers of Olive flounder which will be helpful in population genetic study, linkage mapping construction and selective breeding.Partial genomic library of Olive flounder {Paralichthys olivaceus Temminck et Schlegel) was constructed. Using (AC)₁₀ and (AG)₁₀ as probes 20 microsatellites were identified from 16 positive clones screened, among which 13 were perfect, 5 were imperfect and 2 were compound .By using the home-made software (Repeat Reporter 1.5), the ESTs database of Olive flounder was screened. 265 microsatellite sequences were obtained from 2992 ESTs sequences, about 8.9% in whole database. A total 114 dinucleotide repeats and 88 trinucleotide repeats were found in these 265 microsatellite sequences . (AT)n in dinucleotide repeats and (CTT)n in trinucleotide repeats were the most common.Twenty-eight primer pairs were designed from microsatellites using computer program Primer Premier( Version 5.0) .Of the 19 primers that could amplify PCR product, 16 primers were polymorphic in 30 Olive flounder. In these 16 microsatellite loci, the alleles numbers distributed from 4 to 19. The expected heterozygosity and the PIC values of them distributed from 0.7175 to 0.9613 and 0.663 to 0.94, respectively. These values showed these 16 microsatellite loci were suitable for genetic analysis.