【摘要】 以甘蓝型油菜温敏核不育系SP2S及其近等基因系SP2F为材料,分别采用TCA/丙酮法和Tris-丙酮-酚法提取花蕾总蛋白,并优化电泳体系,对可育和不育花蕾中的蛋白质表达进行差异分析。结果表明:TCA/丙酮法适合于花蕾总蛋白的提取,采用10%浓度的分离胶,上样量为450μg,用pH 4~7的17 cm线性胶条和改良的考马斯亮蓝染色法得到了背景清晰、蛋白点分布均匀且重复性好的电泳凝胶图谱。利用这一实验体系,筛选出SP2S和SP2F之间大量的差异蛋白点。相比SP2F,仅在SP2S中表达的有13个点,上调2倍以上的点有4个,下调2倍以上的点有9个。更多还原

【Abstract】 Using TCA/acetone and Tris-acetone-phenol methods to extract total proteins from fl oral buds respectively, an optimized system of two-dimensional electrophoresis(2-DE) was employed to screen differential protein expression between a temperature-sensitive male sterile line SP2 S and its near isogenic line SP2 F in Brassica napus. The results showed that clear background, well distributed protein spots and reproducible 2-DE maps could be acquired following TCA/acetone extraction, electrophoresis system including 10% SDS-PAGE, 450 μg of sample loading quantity, and 17 cm immobilized p H gradient(IPG) strip with p H 4–7, and modifi ed staining using Coomassie Brilliant Blue. Using this optimized 2-DE system some differential protein spots between SP2 S and SP2 F were found. Compared with SP2 F, 13 spots were specifi cally expressed in SP2 S, 4 of them were up-regulated more than 2-fold and 9 were down-regulated more than 2-fold.更多还原