| The bioinforrnatic and FIASCO(Fast isolation by AFLP sequences containingrepeats) technique were used to development microsatellite markers from Siniperca inthis project.Twenty of microsatellite primers were designed using Primer Premier 5.0software according to the microsatellite DNA sequence (GenBank No:DQ789247～DQ789306) of Siniperca chuatsi isolated by ourselves.Then we used these 20microsatellite loci to analyze genetic diversity and genetic differentiation in the fourwild populations of Siniperca,Siniperca chuatsi, Siniperca kneri, Siniperca obscuraand Siniperca scherzeri.1,A bioinformatic analysis of 304 nucleotide sequences in Siniperca chuatsiGenBank identified 22 sequences containing 30 microsatellites, account for 9.87% ofwholly GenBank database. Cluster analysis indicated that the 16 dinucleotide pairswere the most abundant microsatellites,account for 53.33% of the totalmicrosatellite-containing sequences; 11 trinucleotide repeats and 3 tetranucleotiderepeats were found in these microsatellite sequences,account for 36.66% and 10.01%accordingly. 17 primer pairs were designed from these microsatellite sequences.Among the 17 primer pairs, 13 pairs have amplified products, 6 pairs can achieveclear PCR products by Electrophoresis. Genotype analysis results indicated that: theaverage polymorphism information contents(PIC) were 0.692(ranged:0.567-0.806),were all polymorphic loci.2,This project used the method of FIASCO(Fast Isolation by AFLP SequencesContaining repeats) triumphantly constructed the first microsatellite-enriched libraryof Siniperca chuatsi. One hundred clones were isolated and sequenced from Sinipercachuatsi microsatellite libraries. Sixty clones (60%) contained microsatellites(GenBank Accession Number:DQ789247～DQ789306). We designed 47SSR-primers and synthesized 21 of them,18 polymorphic microsatellite markers werescreened out.3,The Microsatellite DNA technique was used to analyze genetic diversity and genetic differentiation in the four wild populations of Siniperca,Siniperca chuatsi,Siniperca kneri,Siniperca obscura and Siniperca scherzeri.Twenty of microsatelliteprimers were designed using Primer Premier 5.0 software according to themicrosatellite DNA sequence (GenBank No:DQ789247-DQ789306) of Sinipercachuatsi isolated by ourselves. The results showed that:the number of alleles(Na) perlocus ranged from 5-25, the effective alleles(Ne) of each loci were 2.2-22.8.The totalof 293 alleles were found with allele size ranged from 80bp to 301bp. The averageheterozygosity(H) among the four populations of Siniperca chuatsi,Siniperca kneri,Siniperca obscura and Siniperca scherzeri was 0.64,0.58,0.67 and 0.65,respectively.At the 20 microsatellite loci among the. four wild populations,the value ofpolymorphism information content(PIC) was 0.269-0.812,0.375-0.857, 0.258-0.857and 0.332-0.843 respectively, averaged at0.5355,0.5106, 0.551 and 0.5478.Power ofdiscrimination (DP) was 0.32-0.82, 0.50-0.86, 0.36-0.86 and 0.42-0.85, respectively.and probability of paternity exclusion (PPE) was 0.338～0.866, 0.263～0.813,0.263-0.506 and 0.384-0.834, in sequence.The results indicated that the 20microsatellite loci selected were very sensitive and could be used in parentage andkinship determination of further genetic breeding studies in Siniperca.Clusteringanalysis indicated that Siniperca chuatsi and Siniperca scherzeri were classified as thefirst grouped as their genetic distance was relatively lower and kinship was relativelylate. Siniperca kneri and Siniperca obscura could be considered as one branch.Insummary,the UPGMA tree was reflected the evolutionary and breeding historyof thefour wild populations, they belong to the different species in the same genus.These microsatellite markers are useful for genetic background analyzing andgenetic linkage mapping construction of Siniperca chuatsi. Therefore, they provided amass of candidates for comparative genome analysis,marker-assisted selection andanalysis of Quantitative Trait Loci location of Siniperca chuatsi.
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