17β-雌二醇对雄性金鱼卵黄原蛋白的诱导作用

The identification of vitellogenin induction by 17β-estradiol in male Carassius auratus

采用腹腔注射17β-雌二醇的方法诱导雄性金鱼卵黄原蛋白产生,注射浓度为0.05mg·g-1BW,诱导2周后取尾静脉血,离心分离血浆,进行血浆常规聚丙烯酰胺凝胶电泳,通过对卵黄原蛋白特性基团磷、脂和糖蛋白的染色,确定了卵黄原蛋白在电泳图谱上的位置,开发了一种简便、高效的定性卵黄原蛋白的聚丙烯酰胺凝胶电泳法。电泳结果表明,在0.05mg·g-1BW的注射浓度下,2周后17β-雌二醇诱导了雄性金鱼卵黄原蛋白产生,并通过ELISA检测卵黄原蛋白的平均含量为690.2ng·mL-1,与对照组雄性金鱼平均含量为10.7ng·mL-1的差异极显著(P<0.01),比雌性对照组检出量285.5ng·mL-1高1倍多;17β-雌二醇诱导组雄鱼血浆钙离子和血总蛋白含量明显增加。

The vitellogenin of male goldfish (]Carassius auratus) was induced by intraperitoneal injection of 0.05 mg·g^(-1)BW of 17β-estradiol dissolved in ethanol. After two weeks induction, the blood was collected from caudal vessel, then centrifuged at 8 000 r·min^(-1) for 5 min. After plasma native-PAGE, the gel was stained with the method of CBB G250-, phospho-, lipo- and glyco-staining respectively. the location of vitellogenin in the gel was identified. This method provides a practical and effective way of vitellogenin identification. The result of native-PAGE showed that, after the injection of 0.05 mg·g^(-1)BW of 17β-estradiol for two weeks, the male fish can synthesize vitellogenin. The vitellogenin average content in E_2 group was 690.2 ng·mL^(-1) by ELISA, it was significantly different from male control group(P<0.01), whose average content was 10.7 ng·mL^(-1), and it was also much higher than that of the female control group, whose average content was 285.5 ng·mL^(-1). Compared with control male group, the Ca^(2+) and plasma total protein content of the E_2 exposed male group was increased significantly. In control group, the Ca^(2+) contents in plasma of male and female control groups were 1.6 mmol·L^(-1) and 1.7 mmol·L^(-1), respectively. After two-weeks treatment of E_2, Ca^(2+) content of male groups was 50% higher than that of male control group, which was 2.3 mmol·L^(-1). The contents of total protein in the control male and female groups were 3253.1 mg·L^(-1) and 7071.0 mg·L^(-1), respectively. After two-weeks treatment of E_2, the total protein content in plasma was 9616.6 mg·L^(-1), which is 3-folder higher than that of the control male group. So E_2 not only can induce male goldfish to synthesize vitellogenin but also can alter the physical index of fish plasma.