《植物生理学报》 2011, 47(2): 199-204

建立和优化双向电泳分析柱花草根系蛋白谱的方法

陈志坚¹, 严炜^1,2, 孙丽莉^1,2, 刘国道², 廖红¹, 田江^1,*

1 华南农业大学根系生物学研究中心, 广州 510642; 2 中国热带农业科学院热带作物品种资源研究所, 海南儋州 571737

通信作者：田江；E-mail: jtian@scau.edu.cn；Tel: 020-85280156

摘　要：

本研究以柱花草根系为材料, 比较了不同蛋白质提取方法和蛋白上样量等因素对双向电泳方法分析根系蛋白图谱的 影响。结果表明, 在苯酚法提取蛋白中, 加入 0.7 mol•L-1 NaCl 和 20% 乙醇, 并在蛋白沉淀过程中加入 1/10 倍体积 5 mol•L-1 NaCl, 能够有效去除组织样品中的非蛋白成分, 结合使用pH 4~7 范围的IPG胶条, 1 mg根系蛋白可以在双向电泳图谱上分 辨出较多蛋白点, 图谱背景清晰, 该体系适合柱花草根系蛋白质的双向电泳分析。

关键词：双向电泳; 蛋白提取; 柱花草

收稿：2010-10-18   修定：2010-11-24

资助：国家自然科学专项基金项目(309420047)。

Establishment and Optimization of Two Dimensional Electrophoresis System for Analyzing Protein Profiles in Stylo Roots

CHEN Zhi-Jian¹, YAN Wei^1,2, SUN Li-Li^1,2, LIU Guo-Dao², LIAO Hong¹, TIAN Jiang^1,*

¹Root Biology Center, South China Agricultural University, Guangzhou 510642, China; ²Institute of Tropical Crops Genetic Resource, Chinese Academy of Tropical Agriculture Science, Danzhou, Hainan 571737, China

Corresponding author: TIAN Jiang; E-mail: jtian@scau.edu.cn; Tel: 020-85280156

Abstract:

In the study, effects of different methods of protein extraction and loading on analyzing protein profiles of stylo roots were investigated using the two-dimensional electrophoresis (2-DE). The results showed that 2-DE could be optimized by using the modified phenol extraction buffer containing 0.7 mol·L^-1 NaCl and 20% ethanol, the precipitation buffer containing 1/10 volume 5 mol·L^-1 NaCl. Totally, 500 protein spots could be well resolved and detected in the gels when 1 mg protein of roots was separated using pH 4–7 IPG strip. All the results suggested that the method was suitable for the study of 2-DE of stylo roots.

Key words: 2-DE; protein extraction; stylo