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目的 :从大鼠精原细胞、粗线期精母细胞和圆形精子细胞筛选精子发生过程中阶段特异表达基因。 方法 :用牛血清白蛋白梯度沉降法 (STAPUT法 ) ,分别从 9d龄和成年大鼠睾丸中 ,分离出处于减数分裂前、中、后的 3种生精细胞 ,即精原细胞、粗线期精母细胞和圆形精子细胞。运用RNA差异显示方法筛选差示cDNA。 结果 :本实验获得 19个cDNA差示片段 ,并克隆和测序出其中 6个cDNA序列 ,通过非重复序列 (nr)和EST序列同源比较 ,发现 5条cDNA分别与小鼠睾丸、附睾、乳腺、早期胚胎和大鼠卵巢基因高度同源 (88%～ 98% ) ,有一条未知新EST。结论 :差异显示技术是分离与精子发生有关基因的强有力工具。
OBJECTIVE: To screen the stage-specific genes during spermatogenesis from rat spermatogonia, pachytene spermatocytes and round spermatids. METHODS: Three kinds of spermatogenic cells pre-meiosis, middle and post-meiosis were isolated from 9-day-old and adult rat testis by STAPUT method, respectively Line spermatocytes and round sperm cells. Screen differential cDNA using RNA differential display method. Results: Nineteen cDNA fragments were obtained and six of them were cloned and sequenced. By homology comparison between nr and EST sequences, five cDNAs were found to be associated with mouse testis, epididymis, breast , Early embryo and rat ovary gene highly homologous (88% ~ 98%), there is an unknown new EST. Conclusion: Differential display is a powerful tool for isolating genes associated with spermatogenesis.