甘蓝型油菜温敏核不育系SP2S花蕾总蛋白质双向电泳体系的建立及应用  

Establishment and Application of Two-Dimensional Electrophoresis System for Floral Bud Proteome of Temperature-Sensitive Male Sterile Line SP2S in Brassica napus

作　　者：张菁雯[1] 李晓荣[1] 赵燕妮[1] 胡玉梅[1] 徐献锋 于澄宇[1] 

机构地区：[1]西北农林科技大学农学院,旱区作物逆境生物学国家重点实验室,陕西杨凌712100

出　　处：《植物生理学报》2014年第10期1601-1607,共7页Plant Physiology Journal

基　　金：教育部新世纪优秀人才项目(NECT-10-0693);现代农业产业技术体系(ARS-13)

摘　　要：以甘蓝型油菜温敏核不育系SP2ST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450gg，用pH4～7的17cm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。Using TCA/acetone and Tris-acetone-phenol methods to extract total proteins from floral buds respectively, an optimized system of two-dimensional electrophoresis （2-DE） was employed to screen differen- tial protein expression between a temperature-sensitive male sterile line SP2S and its near isogenic line SP2F 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 pg of sample loading quantity, and 17 cm immobilized pH gradient （IPG） strip with pH 4-7, and modified staining using Coomassie Brilliant Blue. Using this optimized 2-DE system some differential protein spots between SP2S and SP2F were found. Compared with SP2F, 13 spots were specifically expressed in SP2S, 4 of them were up-regulated more than 2-fold and 9 were down-regulated more than 2-fold.

关 键 词：甘蓝型油菜 温敏核不育 花蕾 双向电泳 蛋白质 

分 类 号：S565.4[农业科学—作物学]