Isolation And Characterization Of SNPs From Half-smooth Tongue Sole (Cynoglossus Semilaevis)

Half-smooth tongue sole (Cynoglossus semilaevis) is an important economic flatfish and its economic characteristics are mostly controlled by quantitative trait loci (QTL) with complex genetic mechanism. The establishment of high-density genetic map is necessary for the precise location of these QTL to selective breeding and the use of SNP markers is a good solution to it, as SNPs provide higher densities than other markers. But there are not sufficient SNP sites for the construction of genetic mapping or associate study owing to the lack of researches on SNPs of half-smooth tongue sole. This research is aimed to isolate and characterize SNP markers for half-smooth tongue sole.In this study, we used 52 pairs of primers based on the end sequences of fosmid library in female half-smooth tongue sole. By PCR-SSCP assay,24 of the 52 primer pairs appeared polymorphic, which contained 115 SNPs with a frequency of 0.74% after sequencing. Among these SNPs,79 of which are transitions,36 are transversions. As a result, all the detected SNPs are biallelic.Candidate SNPs were amplified with semi-nested allele-specific polymerase chain reaction (AS-PCR), among which 17 are polymorphic, and the result of genotyping were tested by sequencing. AS-PCR with the 17 groups of primers in a wild population showed that the expected and observed heterozygosities of the SNPs ranged from 0.097 to 0.508 and 0.033 to 0.633, and the effective number of alleles ranged from 1.105 to 1.998. The polymorphic information content value (PIC) were calculated, which showed that 8 SNPs are low polymorphic while others are medium. No significant linkage disequilibrium were observed with most loci, as well as three loci did not conform to Hardy-Weinberg equilibrium at the level of P<0.05.