Optimization of hydrostatic pressure, timing, and duration parameters for the induction of tetraploidy in turbot, Scophthalmus maximus

Tetraploid fish are the key source of diploid gametes in polyploid breeding, and they can be induced by disrupting the first mitotic cell cleavage. In this study, the induction protocol of tetraploid by hydrostatic pressure shock and the viability of the tetraploid progenies were investigated in turbot, Scophthalmus maximus. Under water temperature of 14.5 +/- 0.5 degrees C, fertilized diploid zygotes were treated by different combination of timing (65-90 min after fertilization, maf), pressure intensity (60-85 MPa), and duration (2-10 min), respectively. Ploidy level was determined by silver staining of NOR regions, karyotype analysis, and flow cytometry. The optimal protocol for the pressure shock induction was determined to be the combination of timing of 80 maf, 75 MPa of hydrostatic pressure, and 6 min of duration time. Two batches of tetraploid-induced progenies with a total of more than 73,000 morphologically normal larvae were produced. The hatching rates and tetraploidy rates of the two induction groups were 8.97% and 4.09% and 53.33% and 46.67% at 1 day after hatching (dah), respectively. At 150 dah, 1 out of 20 juveniles was identified as tetraploid by karyotype analysis. However, none of tetraploid juveniles was detected by the flow cytometry analysis among the 446 juveniles survived at 365 dah. This preliminary study provides the evidence supporting the large-scale production of tetraploid turbot progenies, thus encouraging further research on the subject.