Construction of Two-Dimensional Polyacrylamide Gel Electrophoresis System for Proteins in Brassica napus

The purpose of this study was to establish a suitable 2-dimensional polyacrylamide gel electrophoresis (2-DE) system for proteomics analysis in Brassica napus. Proteins of rapeseed variety 08127 were separated in gel brands (pH 4–7) using trichloroacetic extraction method under the improved isoelectric focusing (IEF) procedure. A clean 2-DE map was obtained, indicating that the system was effective and suitable for B. napus. This system was validated using young seedling, leaves, and stems from other varieties, and the results were similar to that of variety 08127. The proteomics profiling of seedlings in different ages of B. napus was then analyzed on the basis of this system. In the stems of 30-day-age seedlings, 20 up-regulated and 51 down-regulated proteins were found with differential expression more than 2 folds compared to their corresponding proteins in 15-day-age stems. A few differential protein spots were identified using mass spectrometry method, indicating the effectiveness of the 2-DE polyacrylamide gel electrophoresis system established in this study.

摘要选用油菜品种08127, 采用TCA-丙酮提取方法, pH 4∼7胶条和改进的IEF聚焦程序, 得到较好的蛋白质双向电泳结果, 建立了一种适合甘蓝型油菜的双向电泳体系。利用其他甘蓝型油菜品种的幼苗、叶片和茎中的蛋白检验该实验体系, 都能得到较好的蛋白质双向电泳结果。利用该实验体系比较甘蓝型油菜幼苗不同发育时期蛋白质双向电泳图, 找到二者之间的差异点, 选取部分点成功进行了质谱分析。