Molecular Cytogenetic Analyses In Hybrid Scallop (Chlamys Nobilis ♀×C.farreri♂)

In this study, fertilization of biparental hybr dization of Chlamys nobilis C. farreri was of first investigated by with fluorescence microscopy. Subsequently, traditional karyotype analyses were employed to study the construction of chromosomes of hybrids. GISH (Genomic in situ hybridization) was firstly carried out in the study of crossbreeding research of scallops; meanwhile, we developed a kind of species-specific FISH (Fluorescence in situ Hybridization) probe, used FISH technique to detect their hybrids nature practically, and discussed its prospect of application in shellfish crossbreed research. The results of this study are listed as followings:1. The cytology of the hybrid fertilization and the development of early embryos of C. nobilis C. farreri & were investigated by fluorescence microscopy with HOECHST33258 stained, which testified the feasibility of artificial hybridization between the two interspecific scallops. The results showed that the mature eggs of C. nobilis were at prophase or metaphase of meiosis I . Parts of sperms from C. farreri could bind and penetrate into the eggs from C. nobilis. With the activation of these sperms, the eggs finished their meiotic division and released their first and second polar bodies successively. The male and female pronuclei fused in the central of the eggs after their complete maturation. The fusion of male and female pronuclei and the normal development of early embryos provided the evidence that heterogenous heredity genetically participated in the construction of genome in hybrid.2.Traditional karyotype analyses were undertaken to study the hybrids at their trochophore stage, as well as the parental species. Results showed that through number and morphology of chromosomes, we could easily distinguish the two species. More than 70% hybrids contain karyotype of 2n=35=6m+5sm+11st +13t, NF=46, corresponding to the theoretical expectation of hybrids between the two species. Thisstudy provided further evidences that the combination of different haploid genome from the duplex parental species construct the nucleus genome in most hybrids.3. We investigated the feasibility of GISH technique in the study of shellfish genomic research, and optimized the experimental condition of its practical application. We developed an optimum GISH protocol that might be employed in genomic analysis of scallop hybrids. In this study, GISH was employed to detect different genomic chromosome components and identify donor chromatin in hybrids between C. nobilis and C. Tarreri under the present optimum condition. Results showed that more than 70 percentages of hybrids contain 35 chromosomes. Thereinto, 16 chromosomes were detected with green signals by C. nobilis-genome DNA as probes and FITC-PI as signal detection system; 19 chromosomes were detected with green signals by C. farreri-genome DNA as probes and the same signal detection system, corresponding to the sum of chromosomes from the haploid genomes of C. nobilis (n=16) and C. farreri (n=19). Combining with the previous results, we concluded that by the developmental stage of trochophore, most hybrids still had the karyotype of 2n=6m+5sm+llst+13t with NF=46, according with theoretical expectation of hybrids, which crediblely proved to be a combination of haploid genomes of biparents in hybrids. In this study, we did not detect any abnormal behaviour of chromosomes such as deletion, breakage, recombination and so on. We also didn't detect any haploid metaphases (n=16 or 19) in hybrids.4. We detected a few gynogenetic individuals through the interspecific hybridization between C. nobilis and C. farreri . In this study, we found that 2-4% heterogenous sperms from C. farreri had not penetrated into the eggs form C. nobilis but only binded the membrane of them serving as stimulators. Under the exterior activation, the eggs could finish their first and second meiotic divisions and release two polar bodies successively. Through traditional karyotype analyses, we also detected about 2% unexpe...