二维电泳分离牛精子蛋白的技术研究

二维电泳是蛋白质分离技术并可由于对精子蛋白的分离。本研究旨在通过对双向电泳条件的研究摸索出一种适用于分离牛精子蛋白的二维电泳技术，并利用其对牛精子蛋白进行分离鉴定。在实验中，优化了等电聚焦程序，研究了精子蛋白的不同制备方法、不同上样量、不同胶条长度对电泳结果的影响。结果表明，采用尿素－盐酸胍两步裂解法裂解精子细胞制备蛋白，使用13cm非线性胶条进行蛋白二维电泳，能获得较好的电泳图谱。图谱经二维电泳软件分析，可检测出约800多个蛋白质点，分子量基本分布在10~100KD、等电点约为4~9的区域内。对精子蛋白二维电泳条件的摸索，为后续牛精子X、Y差异蛋白的检测和分析奠定了理论基础。

The study on separating the protein of bovine sperm by two-dimensional gel electrophoresis

Two-dimensional gel electrophoresis a protein separation technique and can be used for separating sperm proteins. The objectives were to develop an optimal two-dimensional gel electrophoresis, and the bovine sperm proteins were isolated with the developed technique. Isoelectric focusing electrophoresis program was optimized, and various sample preparation methods, different loading quantities and strip length were studied. The results indicated that the better proteins map was obtained when the bovine sperm were lysed by two step method of urea-guanidine hydrochloride and the 13cm N-linear IPG strip was used to conduct the isoelectric focusing electrophoresis. More than 800 protein spots could be detected by the imaging analysis system, and the analysis result indicated that molecular weight and isoelectric point of most proteins were from 10 to 100kDa and 4.0 to 9.0 respectively. So it provided a basis for detecting and analyzing the differential proteins of the X, Y sperm.