Chromosome Manipulations In Paralichthys Olivaceus

Chromosome manipulation, mainly including polyploid induction, gynogenesis and androgenesis, is artificial change of the number of a species' chromosome sets through biological, physical or chemical method. As one important part of genetic breeding, chromosome manipulation is an active biotechnology that is addopted to genetic improvement offish species.Artificial induction of polyploid, gynogenesis and androgenesis has been repeatedly reported in fresh water fishes as well as in marine fishes in abroad. However, similar researches in marine enconomic fish species in China were relatively less developed. In this study we report the results of chromosome manipulation experiment in olive flounder Paralichthys olivaceus. Optimum parameters for UV irradiation of sperms, suppression of second polar body release and for suppression of the first cleavage were teset.1. The effects of ultraviolet (UV) irradiation on genetic inactivation of sperms of P. olivaceus and Kareius bicoloratus were examined. Using 2 paralleled UV lamp (20W each), the optimal time of UV irradiation for effective inactivation of P. olivaceus sperms was 60sec when the distance between sperms and the UV lamp was 10cm. The optimum dose for effective inactivation of K. bicoloratus sperms was 40s. Gynogenetic haploids, which was proved by the haploid number of chromosomes, were obtained under these irradiation conditions. The fertilization and hatching rates of the eggs exhibited typical"Hertwig Effect".2. The parameters for induction of gynogenetic diploids in P. olivaceus were optimized. Chromosome diploidy was achieved by suppression of the second polar body release by early cold shocks applied l-10min after fertilization at 0-10癈 with 15-90min durations. The optimal cold shock was at 2-4 that was applied at 3minafter fertilization and lasted for 45min. The highest rate of diploidy was 79.7%. The diploid chromosome number of putative diploid gynogens indicated the effectiveness cold shocks. Large number of gynogenetic diploids were obtained.Meiotic gynogenesis in P. olivaceus was also induced by hydrostatic pressure. Chromosome diploidy was achieved by early pressure shocks applied l-10min after fertilization with pressures range in 40-65MPa that lasted for 2-10min. The optimum pressure shock to induce maximal diploidy (61%) of meiotic gynogenesis was 55MPa pressure that was applied at 3min after fertilization and lasted for 6min.Mitotic gynogenesis was induced by hydrostatic pressure shocks for the first time. Diploid fry were obtained by suppression of the first cleavage by late pressure shocks applied 40-100min after fertilization with pressures range in 45-70MPa that lasted for 2-1 Omui. The highest rate of diploidy ft8%) was achieved by 55MPa pressure that was applied at 3min after fertilization and lasted for 6min.3. A preliminary study of induction of triploid and tetraploid in P. olivaceus was conducted. Triploids and tetraploids were successfully induced by suppression of the second polar body release the first cleavage, respectively. Triploidy and tetraploidy were proved by the maximu number of nucleolus. The inducing rate of triploid and tetraploid was 100% and 87% respectively. Development of triploids at blastula, gastrula and hatching were not significantly different from those of diploid, although triploid developed slowly than diploid did after gastrlation. However, the develoment of tetraploid were worse than diploid and triploid did. The hatching rate of tetraploid was only 1.45%.The result of this study provided bases for the further study of polyploid breeding sex control in P. olivaceus.