【作者】 程晓春；

【导师】 林丹军； 尤永隆；

【作者基本信息】 福建师范大学 ， 发育生物学， 2007， 硕士

【摘要】 本文采用组织学方法观察了江黄颡鱼(Pseudobagrus vachelli)原始生殖细胞(PGCs)的迁移、生殖嵴生成和性腺分化；用免疫组织化学方法，检测了对照组和各实验组鱼苗的脑和性腺组织的芳香化酶活性；以及用RT—PCR的方法，检测芳香化酶基因在脑、性腺、胃肠、肝脏、肾脏的表达情况，探讨温度对芳香化酶基因表达的影响。实验结果显示，1日龄仔鱼PGCs位于鱼体中肠背方的脏壁中胚层中；5日龄时PGCs迁移到背方的腹膜上皮；8日龄时生殖嵴形成；14日龄时原始性腺形成；23日龄性腺开始分化。从孵化后的第10d开始，分别用不同水温(20±0.5℃、24±1.0℃、30±0.5℃、32±0.5℃、34±0.5℃)培育仔鱼达25d。实验结束后统计幼鱼性比和雄性率。对照组(24±1.0℃)和20±0.5℃、32±0.5℃实验组的雌、雄性比接近1：1(分别为1：1.09、1.22：1和0.99：1)；30±0.5℃实验组的为1：4.89，雄性率达83.3±0.7％；34±0.5℃实验组的为2.85：1，雄性率仅为26.4±0.4％。30±0.5℃可使幼鱼性腺分化趋向雄性化，34±0.5℃则使幼鱼性腺分化趋向雌性化。芳香化酶主要定位于鱼的脑组织、卵巢中的卵原细胞和滤泡细胞，以及精巢的间质中。无论是对照组或实验组，在性分化前稚鱼在此时已有微弱的芳香化酶表达。随着性分化进程，芳香化酶的表达逐渐增强。在30±0.5℃高温组，芳香化酶活性较弱，而在34±0.5℃高温组，芳香化酶活性较强。性腺分化后，卵巢的芳香化酶活性比精巢的强。P450aromA的表达仅局限在卵巢；而P450aromB在脑、性腺、胃肠、肾组织中均有表达，其中脑的表达量较高。在卵巢中P450aromA的表达量是P450aromB的约2.2倍。更多还原

【Abstract】 In this paper, the migration of primordial germ cells (PGCs), formation of genital ridges and gonad sex differentiation of teleost, Pelteobagrus vachelli was investigated using histological methods, and the distribution of P450 aromatase in some tissues of the fish was examined by immunohistochemistry, too. Also in the paper, the effect of temperature upon expression of P450 aromatase gene was studied by RT-PCR techniques.The results were as follows:On day 1, the PGCs were at the splanchnic mesoderm above the gut. On day 5, PGCs migrated to the peritoneal epithelium. The genital ridge was formed on day 8, and there was no PGCs in it. The primordial gonads were formed on day 14. On day 23, the primordial gonads differentiated into ovaries or testes. From day 10 on, the larvae were cultivated in different water temperatures (20±0.5℃, 24±1.0℃, 30±0.5℃, 34±0.5℃) for 25 days. The control group was cultivated at natural water temperature (24±1.0℃). The sex ratio and percentage of male larvae were calculated after the experiment. At natural water temperature (24±1.0℃) and lower water temperature (20±0.5℃), the sex ratio was approximately 1:1 (1:1.09 and 1.22:1 respectively). However in the group cultivated at 30±0.5℃, the sex ratio was 1:4.89 and the percentage of male was 83.3±0.7%. The sex ratio was 2.85:1 and the percentage of male rate was only 26.4±0.4% in the group cultivated at 34±0.5℃. These results suggested that water at 30±0.5℃influenced the larval gonad towards differentiating into a testis, and water at 34±0.5℃gave the opposite result, i.e. the gonad differentiated into an ovary. This experiment showed that the sex determination of P. vachelli was temperature-dependent sex determination.The P450 aromatase was mainly orientated in the brain, oogonia and follicular cells in the ovaries, and interstitial tissue in the testes. Before gonad sex differentiation, there was weak P450 aromatase activity in larvae of both control and experimental groups. With the process of gonad sex differentiation, the activity of P450 aromatase increased. In the group cultivated at 30±0.5℃, P450 aromatase activity was lower than that in the group cultivated at 34±0.5℃. In the larvae with completion of gonad differentiation, P450 aromatase activity in the ovaries was higher than that in the testes. P450aromA mRNA was detected only in ovaries. Whereas P450aromB mRNA was identified in brain, gonads, stomach, intestines and kidney, with more amount in brain. In the ovaries, the amount of P450aromA mRNA was about 2.2 times as much as P450aromB mRNA.更多还原