ABSTRACT
Three methods of triploidy (3N) induction were tested in diploid (2N) oysters Crassostrea gigas: two chemical methods cytochalasin-B (CB) and 6-dimethylaminopurine (6-DMAP), and one physical method with temperature shock. The objective was to evaluate the triploidy induction technology using flow cytometry as a tool to check the results of induction. The experiments were performed in separate and a seawater temperature in the tanks was maintained at 25 C for all experiments. In the experiment I, the efficacy of triploidy induction was evaluated using CB (0.5 mg L-1) and 6-DMAP (390 mols L-1). In the experiment II, the efficiency of triploidy induction was tested using CB (0.5 mg L-1) and 6-DMAP (450 mols L-1). In the experiment III, the efficiency of triploidy induction was evaluated using CB (0.5 mg L-1) and temperature shock (25 to 36 C). In all three experiments, viable triploid larvae were obtained. However, in the experiments I and II (with chemical methods), high mortality of larvae was observed, especially for the treatment CB. From these results, it is suggested the replacement of CB by other methods of triploidy induction, due to its high cost and high toxicity to humans and to the environment.
Três métodos de indução à triploidia (3N) foram testados em ostras diplóides (2N) (Crassostrea gigas); dois métodos químicos, citocalasina-B (CB) e 6-dimetilaminopurina (6-DMAP), e um método físico, com choque de temperatura. O objetivo foi avaliar a tecnologia de indução à triploidia, utilizando a técnica de citometria de fluxo como ferramenta para verificação dos resultados de indução. Os experimentos foram realizados em separado, sendo que a temperatura da água do mar foi mantida em 25 C em todos os tanques. No experimento I, foi avaliada a eficácia da indução à triploidia com CB (0,5 mg L-1) e 6-DMAP (390 mols L-1). No experimento II, foi testada a eficiência da indução à triploidia com CB (0,5 mg L-1) e 6-DMAP (450 mols L-1). No experimento III, foi avaliada a eficiência da indução à triploidia com CB (0,5 mg L-1) e choque de temperatura (25-36 C). Nos três experimentos, foram obtidas larvas triplóides viáveis. Entretanto, nos experimentos I e II (com métodos químicos), observou-se elevada mortalidade das larvas, especialmente para o tratamento CB. A partir destes resultados, a substituição de CB por outros métodos de indução à triploidia é sugerida, devido ao seu elevado custo e elevada toxicidade para os seres humanos e para o meio ambiente.
Subject(s)
Animals , Adenine/administration & dosage , Cytochalasin B/administration & dosage , Crassostrea/anatomy & histology , Crassostrea/drug effects , TriploidyABSTRACT
Three methods of triploidy (3N) induction were tested in diploid (2N) oysters Crassostrea gigas: two chemical methods cytochalasin-B (CB) and 6-dimethylaminopurine (6-DMAP), and one physical method with temperature shock. The objective was to evaluate the triploidy induction technology using flow cytometry as a tool to check the results of induction. The experiments were performed in separate and a seawater temperature in the tanks was maintained at 25 C for all experiments. In the experiment I, the efficacy of triploidy induction was evaluated using CB (0.5 mg L-1) and 6-DMAP (390 mols L-1). In the experiment II, the efficiency of triploidy induction was tested using CB (0.5 mg L-1) and 6-DMAP (450 mols L-1). In the experiment III, the efficiency of triploidy induction was evaluated using CB (0.5 mg L-1) and temperature shock (25 to 36 C). In all three experiments, viable triploid larvae were obtained. However, in the experiments I and II (with chemical methods), high mortality of larvae was observed, especially for the treatment CB. From these results, it is suggested the replacement of CB by other methods of triploidy induction, due to its high cost and high toxicity to humans and to the environment.(AU)
Três métodos de indução à triploidia (3N) foram testados em ostras diplóides (2N) (Crassostrea gigas); dois métodos químicos, citocalasina-B (CB) e 6-dimetilaminopurina (6-DMAP), e um método físico, com choque de temperatura. O objetivo foi avaliar a tecnologia de indução à triploidia, utilizando a técnica de citometria de fluxo como ferramenta para verificação dos resultados de indução. Os experimentos foram realizados em separado, sendo que a temperatura da água do mar foi mantida em 25 C em todos os tanques. No experimento I, foi avaliada a eficácia da indução à triploidia com CB (0,5 mg L-1) e 6-DMAP (390 mols L-1). No experimento II, foi testada a eficiência da indução à triploidia com CB (0,5 mg L-1) e 6-DMAP (450 mols L-1). No experimento III, foi avaliada a eficiência da indução à triploidia com CB (0,5 mg L-1) e choque de temperatura (25-36 C). Nos três experimentos, foram obtidas larvas triplóides viáveis. Entretanto, nos experimentos I e II (com métodos químicos), observou-se elevada mortalidade das larvas, especialmente para o tratamento CB. A partir destes resultados, a substituição de CB por outros métodos de indução à triploidia é sugerida, devido ao seu elevado custo e elevada toxicidade para os seres humanos e para o meio ambiente.(AU)
Subject(s)
Animals , Crassostrea/anatomy & histology , Crassostrea/drug effects , Adenine/administration & dosage , Cytochalasin B/administration & dosage , TriploidyABSTRACT
The current study examined the protective effects of l-glutamine and cytochalasin B during vitrification of immature bovine oocytes. Oocyte vitrification solution (PBS supplemented with 10% FCS, 25% EG, 25% DMSO and 0.5 m trehalose) was the vitrification control. Treatments were the addition of 7 µg/ml cytochalasin B, 80 mm glutamine or both cytochalasin and glutaminine for 30 s. After warming, oocytes were matured in vitro for 24 h, fixed and stained with Hoechst (33342) for nuclear maturation evaluation. L-glutamine improved the vitrified/warmed immature bovine oocytes viability (32.8%), increasing the nuclear maturation rates compared to other treatments and the no treatment vitrified control (17.4%). There was, however, no effect of cytochalasin B on in vitro maturation (14.4%).