|Other Abstract||The common carp (Cyprinus carpio) and bighead carp (Aristichthys nobilis) are two widely distributed fish species and have been intensively cultured in China. In the past a few decades, fish resources in Chinese inland waters have been severely influenced by over-fishing and environmental deterioration. Furthermore, lack of scientific management system of artificial breeding has led to the decline of germplasm resources of cyprinid fishes including common carp and bighead carp. In the present study, we developed Type I markers and analyzed population genetic in the two carps, and compared various genotypes associated with growth traits. We aimed to provide theoretical and applied bases for further studies on genetic resource conservation and marker-assisted selective breeding in common carp and bighead carp. The main results of the dissertation are shown as follows:
1. A total of 18840 ESTs (Expressed Sequence Tag) of common carp were downloaded from public databases and analyzed by bioinformatics methods. In total, about 5.5% of these ESTs had SSRs (Simple Sequence Repeats) inside. After clustering and assemblying, 903 unique ESTs were identified. The abundance of di-, tri-, tetra-, and penta-nucleotide motifs among the ESTs is 37.18%, 30.81%, 20.36%, and 11.71%, respectively. The rates of polymorphic EST-SSRs were 17% in crucian carp (Carassius auratus), and 5% in silver carp (Hypophthalmichthys molitrix), respectively when cross-species amplificatons were tested. 470 pairs of EST-SSR primers were designed for these novel EST-SSRs in order to provide sufficient polymorphic markers for population genetic studies and genome mapping in common carp and its closely related fishes.
2. Population genetic diversity of the common carp (ten wild populations and one cultivated population) was investigated based on 10 polymorphic EST-SSR loci. On the whole, the genetic diversity of wild common carp population is high, and the genetic diversity in Heilongjiang population is lower than in Yangzi River populations. The Fst value (0.10) showed that common carp had moderate genetic differentiation, and the gene flow analyses indicated that the genetic exchange among populations is not difficult. There were no significant relations between genetic distances and geographical distributions of the eleven populations based on NJ (Neighbor-Joining) clustering and SOM (Self-Organized Map). These results indicate that EST-SSRs in common carp are non-neutral molecular markers, and they would be helpful to investigate the functional diversity of genes and adaptability of common carp populations from major waters in China.
3. Four polymorphic EST-SSR loci were used to explore the relationship between genotypes and growth traits of common carp. As the results, the effects of the four EST-SSR loci on one or more the growth traits (body length, weight, and relative fatness) were evident (P<0.05). These results suggest these EST-SSRs loci may be associated with the growth traits of common carp, and is necessary to be further studied.
4. Gene cloning and searching for genic markers were performed for the Leptin receptor gene (Lepr) in bighead carp. Homology comparisons showed that compared with mammals, the Lepr gene in fish is relatively conserved. One SSR and two SNP (single nucleotide polymorphism) loci were identified in bighead carp population by PCR-SSCP (single strand conformation polymorphism) analysis and DNA direct sequencing. At the SSR locus, the difference in genetic diversity between breeding populations and wild population was not significant. The associations between Lepr gene polymorphisms and growth traits were estimated with the general linear model. The result showed that there were no significant effects on the growth trait at the genic SSR locus in Lepr. At the SNP loci, the individuals with genotype Aa had a larger body height than those with genotype AA and aa (P<0.05); the individuals with genotype Bb had a larger body height and head length than genotype bb (P<0.05); as a dominant allele in the population, the individuals with allele G had a larger body length, body height, head length, body weight and relative fatness than those with allele A (P<0.05). These data showed that the genotypes of SNP loci in Lepr gene were significantly associated with some of the growth traits in bighead carp and could serve as potential candidate gene in further studies of marker-assisted breeding.|