Isolation Sex Specific Microsatellite Marker And Genetic Analysis In F₁Family Of The Rock Bream Fish Oplegnathus Fasciatus

The rock bream fish (Oplegnathus fasciatus) belongs to the family Oplegnathidae andmainly distributed in the Yellow Sea, the East China Sea and Taiwan channel. It is one ofthe most economical important marine fish in China. Due to its high commercial andornamental value, the artificial culture of rock bream has increased significantly in the lastdecade. In order to sustainable development, wild resource protection and benefit themolecular breeding of fish species, the three items were studied in this paper including:(1)Isolation, cloning sex-linkage microsatellite DNA marker and development genetic sexidentification technique;(2) Isolation of polymorphic microsatellite markers;(3)Segregation pattern of AFLP marker in F1family of the rock bream (Oplegnathusfasciatus).(1) Isolation of polymorphic microsatellite markers:One repeat enriched genomic libraries (the repeat units was CA) was constructed byFIASCO (fast isolation by AFLP of sequences containing repeats) in the Oplegnathusfasciatus.60sequences were obtained, of which49contained microsatellite repeats.17pairs of primers were designed successfully, and15microsatellite loci were proved to bepolymorphic. The number of alleles per polymorphic locus ranged from4to9in a sampleof30individuals. Observed and expected heterozygosities per locus varied from0.267to0.767and from0.395to0.859, respectively. Eleven of15microsatellite loci wereconformed to Hardy–Weinberg equilibrium. No significant linkage disequilibrium betweenpairs of loci was detected. The present microsatellite markers could be benefit for theevaluation of genetic resource and the construction of genetic linkage map in O. fasciatus.(2) Isolation, cloning sex-linkage microsatellite DNA marker and development geneticsex identification technique:A sex-linkage microsatellite DNA marker of Oplegnathus fasciatus was isolated. In thisstudy, a male specific allelic gene was detected and specific primers were designedaccording the allele. Subsequently, the simple and fast technique for genetic sexidentification was developed. This method has the advantage of accurate, sensitive andreliable. Such molecular markers can be used for sex identification at an early life stage andare therefore useful for the selective breeding. In addition, our results provide new insightinto the mechanism of sex determination and evolution of sex chromosomes in this species.(3)Segregation pattern of AFLP marker in F1family of the rock bream (Oplegnathusfasciatus). In this study, the segregation pattern of AFLP (Amplified fragment lengthpolymorphism) are investigated using14selective primer combinations. A total of528lociwere produced, of which227loci were polymorphic (42.9%), and301loci weremonomorphic. Among227segregating loci,138loci segregated in Mendelian ratio while89loci segregated in distortion estimated by Chi-square test (P=0.05). There were118and109loci segregating in the ratios of3:1and1:1, respectively. The number od locisegregated in Mendelian ratio were47for3:1and91for1:1, respectively. Effectivenumber of alleles, shannon genetic diversity index and Nei’s gene diversity index werebetween1.1146to1.4326, between0.1393to0.1919, and between0.0836to0.1116,respectively.