油菜种子蛋白质提取方法研究

Study on Protein Extraction Methods for Seeds of Rapeseed

通过对比改进的Tris-HCl法、酚提取-甲醇/醋酸铵沉淀法、TCA/丙酮沉淀法和DTT/丙酮法4种方法提取油菜未成熟种子蛋白质,以期找到合适的蛋白质提取方法。结果表明,TCA/丙酮沉淀法和DTT/丙酮法所得蛋白质沉淀较多,颜色亮白,效果较好。进一步通过定量分析及双向电泳图谱的比较分析表明,TCA/丙酮沉淀法在2-DE图谱上蛋白质点较多,分布均匀,图像清晰,而DTT/丙酮法相对于本实验中TCA/丙酮法检测到较少的蛋白质点外,2-DE图谱上有明显的纵条纹。因此TCA/丙酮沉淀法为提取油菜种子中蛋白质的最优方法。

To screen optimal protein extraction method for immature seeds of rapeseed,four methods,the improved Tris-HCl method,phenol extraction -methanol/ammonium acetate precipitation,TCA/acetone precipitation and DTT/acetone method,were compared. The results showed that the TCA/acetone precipitation method and DTT/acetone method could gain more protein precipitation,more bright white color,and the effect was better. Further quantitative analysis and two-dimensional electrophoresis comparative analysis showed that TCA/acetone precipitation method could gain more protein points on 2-DE map,and the distribution was uniform,the image was clear,while DTT/acetone method detected less pro-tein points,and the 2-DE map had obvious vertical stripes. So TCA/acetone precipitation method was the optimal protein extraction method for seeds of rapeseed.