摘要
目的运用蛋白质组学方法,寻找大鼠精子发生相关蛋白。方法用重力沉降法(STAPUT法)从9d龄雄性大鼠睾丸中分离出A型精原细胞,从成年的雄性大鼠睾丸中分离出粗线期精母细胞、圆形精子细胞。分别提取这3种细胞的总蛋白,进行双向电泳。对所得到的双向电泳图谱进行分析,找出差异蛋白。对挑选出的差异蛋白做质谱分析。结果发现磷脂酰乙醇胺结合蛋白(RKIP)在这3种不同细胞的表达上有明显差异。在双向电泳图谱中,RKIP在圆形精子细胞中表达量较高,并且形成分子量相同但等电点不同的并排4个蛋白质点；而在A型精原细胞和粗线期精母细胞的双向电泳图中,RKIP仅为一个单独的蛋白质点。结论RKIP在精子发生过程中有明显的差异表达,可能对精子发生起重要调节作用。
Objective The two-dimensional electrophoresis（2-D） method was used for screening the stagespecific expression proteins during rat spermatogenesis. Methods The highly enriched spermatogonia, pachytene spermatcoytes and round spermatids were isolated by STAPUT method （sedimentation velocity at unit gravity, with 2%-4% BSA gradient in DMEM/F12 medium） to get the total proteins from the three kinds of cells. The difference of protein expression in the three kinds of cells was respectively analyzed by two-dimensional electrophoresis. Results By means of two- dimensional electrophoresis, The raf kinase inhibitor protein （RKIP） was exclusively detected.The RKIP presented four protein dots with the same molecular weight but different isoelectric point in the round spermatids 2D map, only one protein dot in the spermatogonia and pachytene spermatcoytes2D map respectively. Conclusion These results suggested that RKIP is a stage-specifical expression protein and may play an important role in spermatogenesis.
出处
《中国男科学杂志》
CAS
CSCD
2006年第3期38-42,共5页
Chinese Journal of Andrology
