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当前位置: 首页 > 文章 > 油菜叶片总蛋白质双向电泳样品制备方法的改进 生物技术通报 2010,2010 (5) 93-96+110
Position: Home > Articles > Improvement of Preparation Methods of Two-Dimensional Eleteophoretic Samples of Proteome from Brassica Leaves Biotechnology Bulletin 2010,2010 (5) 93-96+110

油菜叶片总蛋白质双向电泳样品制备方法的改进

作  者：
孔芳;蒋金金;吴磊;葛才林;王幼平
单  位：
扬州大学生物科学与技术学院
关键词：
油菜;叶片总蛋白;双向电泳;图谱
摘  要：
以甘蓝型油菜"扬油6号"的叶片为试验材料,分别采用传统的TCA/Acetone(三氯乙酸/丙酮沉淀法)和改进的PEG(polyethylene glycol)分步提取法提取叶片可溶性总蛋白,并利用条件一致的蛋白质双向电泳体系进行比较。TCA/Acetone法提取的蛋白质双向电泳图谱背景中由于高丰度"housekeeping"结构蛋白的存在,特别是叶片中参与光合作用的Rubisco蛋白的干扰,图谱中低丰度调控蛋白受到了高度覆盖和遮蔽现象,影响双向电泳图谱的质量。而PEG分步提取法提取的蛋白质样品,可以剔除Rubisco蛋白,使获得的双向电泳图谱清晰,无斑点间的遮蔽现象,为油菜叶片蛋白质组定量和定性分析提供了丰富的信息。
译  名：
Improvement of Preparation Methods of Two-Dimensional Eleteophoretic Samples of Proteome from Brassica Leaves
作  者：
Kong Fang Jiang Jinjin Wu Lei Ge Cailin Wang Youping(College of Bioscience and Biotechnology,Yangzhou University,Yangzhou 225009)
关键词：
Brassica Proteome from leaves Two-Dimensional electrophoresis Map
摘  要：
With leaves of Brassica napus L.cv."YangYou 6" used as experimental materials,the proteins were extracted from leaves by TCA/Acetone precipitation and differential PEG(Polyethylene glycol)precipitation method and compared by two-dimensional electrophoresis system with uniform conditions of protein.The background noise of two-dimensional eleteophoresis map of protein extracted by TCA/Acetone method was great,and clarity was poor.This was partially attributed to the existence of high-abundant proteins,such as ribulose-1,5-bisphosphate carboxylase/oxygenase(Rubisco) in leaves.They engaged a large proportion of the whole-cell proteins and thus preventd low-abundant proteins from being up-taken by immobilized pH gradient(IPG) strip,consequently making the latter poorly detectable by 2-DE.So they formed high coverage,shading on the other protein spots and affected the accurate expression of map information.However,through depletion of the high-abundant protein Rubisco during differential PEG precipitation method,the background noise of electrophoresis map was little,and PEG fractionation improved detection of low-abundant proteins,there was no shading phenomenon among spots.Therefore it provided abundant information for quantitative and qualitative analyses on the proteomes from Brassica leaves.

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