A reference PLOT for Accurately Predicting the percentage of triploid larvae in spawns
Markets for triploid (three sets of chromosomes) fishes and bivalves are based on the assumption that triploids, offer some advantage over diploids, such as population management, improving growth rates, and increasing survival. Triploid induction treatments, however, do not assure 100% triploid individuals. Estimating the efficacy of these treatments would enable managers to stock fry from spawns exhibiting high triploidy, thereby conserving aquaculture resources such as space, time, labor, and feed. The objective of this study was to generate a reference plot based on ploidy data derived via flow cytometry to obtain accurate early estimates of triploid percentages within grass carp (Ctenopharyngodon idella) spawns.
At 3 days post-hatch, larvae were mechanically dissociated into single-cell suspensions, and nuclear DNA was stained with a propidium iodide solution. Following flow cytometric analyses of larval individuals, aliquots of diploid and triploid cell suspensions were pooled to generate 16 levels of known ploidy, ranging from 0% to 100%. In ten trials using either 20 or 50 individuals, the observed triploidy percentages estimated by flow cytometry histogram values were consistently lower than the known, actual ploidy values (Fig. 1). This is likely due to fewer cells per individual and larger nuclei in triploids. The DNA distribution data were statistically analyzed and fit to the curves whereby the actual triploidy value can be predicted with a formula generated to match the curve (Fig. 2). Thus, a reference graph with its corresponding formula can be used to predict, within 95% confidence limits, the ploidy status of fry in grass carp spawns. This method was used with black carp (Mylopharyngodon piceus) and white crappie (Pomoxis annularis) x black crappie (P. nigromaculatus). This prediction method can be applied with any aquaculture species induced for triploidy.