ISOLATION AND GENETIC ANALYSIS OF MICROSATELLITE MARKERS FOR LARIMICHTHYS CROCEA

Abstract

We constructed an(AC)n-microsatellite-enriched library for large yellow croaker Larimichthys crocea(Richardson 1846) using the method of FIASCO.Ninety clones were randomly selected for further colony amplification.The rate of positive clones reached to 66.7%,and 56 of the 60 positive clones sequences contained one or more SSR.By the analysis of repeat motif,dinucleotide repeats were the most dominant(91.1%),followed by trinucleotide repeats(8.9%).Among dinucleotides,(AC)n repeats were the most frequent(94.1%).Using Weber’s classification rules,the sequences were divided into three categories,the percentage of perfect repeat sequences,imperfect repeat sequences,and compound repeat sequences was 75.0%,8.9%,and 16.1,respectively.Fifty two microsatellites primer pairs were designed and synthesized.The result showed that 35 SSRs were polymorphic in 46 F1 generation individuals.Twenty eight(80.0%) loci exhibited a segregation ratio of 1∶1∶1∶1(AB × CD/AB × AC),serving as the most useful markers segregating in co-dominant fashion.Six loci exhibited a segregation ratio of 1∶1,and one loci exhibited a segregation of 1∶2∶1.Polymorphism analysis showed that 32 SSRs were consisted with Mendelian segregation ratio and could be used to construct the linkage map,while the other 3 SSRs(LYC0137,LYC0139,LYC0152) were departure from the expected Mendelian segregation patterns.These SSRs should be available for genetic linkage mapping,population genetics studies,molecular evolution and phylogenetic study of large yellow croaker.