奶牛乳腺组织蛋白质样品的制备及2-DE图谱分析

本研究以奶牛乳腺组织为对象,采用了普通离心、超速离心、2-D clean-up kit和TCA/丙酮沉淀方法制备蛋白质样品,经二维凝胶电泳(2-DE)获得蛋白表达图谱。根据凝胶图谱上蛋白质斑点判断图谱的质量,并运用PDQuest7.4软件分析图谱。结果显示:普通离心法制备的蛋白样品的凝胶图谱蛋白质斑点较为清晰,但横向条纹较多;超速离心法纯化蛋白样品的图谱,蛋白斑点清晰且可检测到较多的斑点;TCA/丙酮沉淀和2-D clean-up kit方法纯化蛋白样品的凝胶图谱条纹减少,蛋白斑点清晰,但可检测到的斑点数量有所减少。研究表明,超速离心方法纯化的蛋白质样品适合建立奶牛乳腺组织的蛋白质表达图谱。

Preparation of Protein Samples from Mammary Tissues and Its 2-DE Analysis in Dairy Cow

The mammary gland is a complex organ,and associated with immune factors and secretion that play a vital role in protecting the mammary gland from infection or inflammation.Centrifugation,ultracentrifugation,2-D clean-up kit and TCA/acetone methods were used to prepare samples and corresponding 2-DE maps were compared by PDQuest7.4 software.The results showed that samples prepared by ultracentrifugation resulted in an efficient concentration,desalting and removing non-protein,its 2-DE map has showed clear spots with less background than the others.Precipitation with 2-D clean-up kit and TCA/acetone were also efficient desalting and removing non-protein,these gel maps have strong spots,but the protein spots less than that of ultracentrifugation method.The results indicated that ultracentrifugation method was a efficient sample preparation of mammary gland for proteomic analysis.