Research Of Triploid Induction By Hypotonic Treatment In Scallop Patinopecten Yessoensis

Research progress of triploid breeding of shellfish was reviewed in this paper. Methods and basic principle of triploid induction, influential factors in induction and the biological characteristics of the triploid mollusk were discussed. Problems in mollusk triploid breeding were analyzed and prospects were viewed. A new method of polyploid induction was developed and used in scallop Patinopecten yessoensis for triploid induction by inhibiting polar bodyⅡ(PB2)releasing with hypotonic treatment. The suitable induction parameters were studied. Triploidy induction and larvae nursery were conducted with observation and analysis on the growth, survival and variation of ploidy during the stage of larval development and spat. Also the ultrastructure of fertilized eggs with hypotonic treatment were observed with transmission electron microscope(TEM).The main contents consist of the following three parts.1. Study on the suitable induction parameters of triploid Patinopecten yessoensis with hypotonic treatment.In order to optimize the induction parameters, including the hypotonic salinity, the beginning time of treatment and the duration time, different hypotonic treatments (Salinity of 8, 10, 12, 14, 16and 18; when 30% of fertilized eggs released PB1, when 50% of fertilized eggs released PB1, when the first PB2 appeared; Duration time of 10min, 15min, 20min, 25min)were carried by inhibiting polar bodyⅡ(PB2)releasing. Results showed when the fertilized eggs were treated 20min with a hypotonic treatment of 12 salinity at the time when the first PB2 was observed, can yield that the highest triploidy rate which was up to 84.74%, and the average triplody rate was (81.12±3.82)%.2. Nursery and spat cultivation in triploidsTriploid Patinopecten yessoensis induced by hypotonic treatment using the optimized induction parameters, was nursed during larvae stage and growed up to 12 month-old. The growth, survival and ploidy of triploid larvae were studied in Haiyang. The cleavage rate and hatching rate of treatment group was 39.03% and 42.91%, and the control groups was 48.63% and 58.25%。The induced triploid larvae appeared much advantageous in growth, especially when the umbo-stage larva (P<0.001), while the advantage in growth of eyespot larvae was lower(P<0.05). The survival rate was lower than that of the diploid controls and the relative survival rate dropped gradually. The triploidy rate of initial D-shape larvae was 73.86%, and it dropped to 56.47% after 13 days of larvae breeding. Another triploid breeding was carried in Zhangzidao where the survivable triploid juvenile were successfully produced. The triploidy rate in period of lavae, spat, juvenile of 5months, 8 months and 12 months old was 55.81%, 35.00%, 13.33%, 8% and 8%, respectively. The rate of 2n/3n mosaic were determined in juvenile of 5months and juvenile of 8 months by folwcytometer, which was 3.33% and 2.00%, respectively. The triploid juvenile didn't show advantageous in growth than diploids probably due to the deficiency in amount.3. The ultrastructure observation of fertilized eggs in Patinopecten yessoensis induced with hypotonic treatment.The change of fertilized eggs'ultrastructure before and after hypotonic treatment were observed by TEM and studied comparelly. Result showed that the ultrastricture of fertilized eggs changed largely under the influence of hypotonic treatment: Egg membrane was destroyed. Plasma membrane crimpled and dissolved partly. Enchylema was sparse. The yolks were crimpled. Mitochondrias expanded, appeared many vacuoles and even got membrane destroyed. Golgi apparatus appeared some vacuoles. Ribosomes accreted on the rough endoplasmic reticulum fell off. The fertilized eggs repaired itself for a certain extent after being dipped in the natural seawater for a period of time: Plasma membrane got more level up. Cytoplasm was a little plumpish. The yolks had smooth and inerratic figure. Mitochondrias were natural. The results indicate that hypotonic treatment breaks down cell organs such as mitochondria etc.; it causes the metabolic disturbance of fertilized eggs. This is one of the reasons how the low cleavage rate and hatching rate of the triploid Patinopecten yessoensis.