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[1]雍彬,姜玉松,樊汶樵,等.不同方法对甘薯块根可溶性蛋白的提取效果[J].应用与环境生物学报,2014,20(02):211-216.[doi:10.3724/SP.J.1145.2014.00211]
 YONG Bin,ZHANG Yizheng,FAN Wenqiao,et al.Effect study of different methods to extract protein from sweet potato tuberous roots[J].Chinese Journal of Applied & Environmental Biology,2014,20(02):211-216.[doi:10.3724/SP.J.1145.2014.00211]
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不同方法对甘薯块根可溶性蛋白的提取效果()
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《应用与环境生物学报》[ISSN:1006-687X/CN:51-1482/Q]

卷:
20卷
期数:
2014年02期
页码:
211-216
栏目:
研究论文
出版日期:
2014-04-25

文章信息/Info

Title:
Effect study of different methods to extract protein from sweet potato tuberous roots
作者:
雍彬姜玉松樊汶樵张义正
1四川师范大学生命科学学院 成都 610101 2重庆文理学院林学与生命科学学院 重庆 402168 3四川大学生命科学学院四川省分子生物学及生物技术重点实验室 成都 610064
Author(s):
YONG Bin ZHANG Yizheng FAN Wenqiao JIANG Yusong
1College of Life Science, Sichuan Normal University, Chengdu 610101, China 2Institute of Forestry and Life Sciences, Chongqing University of Arts and Sciences, Yongchuan 402168, China 3Sichuan Key Laboratory of Molecular Biology & Biotechnology, School of Life Sciences, Sichuan University, Chengdu 610064, China
关键词:
甘薯块根蛋白质制备SDS-PAGE双向电泳蛋白质组学
Keywords:
sweet potato tuberous root two dimensional electrophoresis (2-DE) protein extraction methods SDS-PAGE proteomics
分类号:
TQ93 : S531.01
DOI:
10.3724/SP.J.1145.2014.00211
文献标志码:
A
摘要:
为建立一套适宜甘薯块根蛋白质组学分析的双向电泳体系(2-DE),采用匀浆法、丙酮沉淀法、TCA-丙酮沉淀法和酚提取法来制备甘薯块根可溶性总蛋白,并分别通过蛋白质定量、SDS-PAGE和2-DE技术来评估这4种蛋白制备方法提取甘薯块根蛋白质的优劣. 结果显示,每0.5 g鲜薯块根,经Bradford方法测定蛋白质的得率分别为1.6 mg、2.4 mg、1.2 mg、1.9 mg. SDS-PAGE的结果表明,匀浆法、丙酮法制备的蛋白质分离到的条带较少,且在约Mr 20 × 103处的储藏蛋白含量较高;TCA-丙酮法和酚提取法制备的蛋白质分离到的条带则明显多于前两种方法,且Mr 20 × 103处储藏蛋白的相对含量也明显减少. 2-DE结果显示,匀浆法制备的蛋白质在进行双向电泳分析时,几乎不能完成聚焦过程,仅在凝胶的碱性端出现少量的蛋白质斑点,丙酮法和TCA-丙酮法要优于匀浆法,但是蛋白质斑点数量仍然较少,酚提取法提取的蛋白质在进行双向电泳时分离到的蛋白质斑点数最多,为1 120个,且在凝胶上均匀分布,背景清晰. 综上所述,酚提取法是适宜甘薯块根蛋白质双向电泳样品制备的最佳方法.
Abstract:
In order to establish a two-dimensional electrophoresis system for the proteomics study of sweet potato tuberous roots, this study compared four protein extraction methods (homogenate method, acetone precipitation method, TCA- acetone precipitation method and phenol method). Then we evaluated these four protein preparation methods through different parameters, such as protein yield, SDS-PAGE and 2-DE techniques. In this experiment, 1.6 mg, 2.4 mg, 1.2 mg and1.9 mg soluble protein was yielded from 0.5 g fresh potato tuber using these 4 protein extraction methods, respectively. The SDS-PAGE results showed that the homogenate method and acetone precipitation method generated only few protein bands with high storage protein content (about Mr 20 × 103). By contrast, the TCA- acetone precipitation method and phenol method could separate more protein bands, with much lower band of storage protein. On 2-DE gels, the homogenization method could hardly complete the IEF, with only a few protein spots appearing in its alkaline end. The results of acetone method and TCA-acetone precipitation method were obviously better than homogenization method, but the number of protein was still less than phenol method. The number of protein spot obtained by phenol method was the most, and these protein spots were evenly distributed in the 2-DE gel. In summary, the phenol extraction method is suitable for sample preparation of sweet potato tuber for two-dimensional electrophoresis.

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备注/Memo

备注/Memo:
国家科技支撑计划资助项目(2007BAD78B03)、四川省教育厅重点项目(13ZA0146)和四川师范大学重点培育项目(14py03)资助
更新日期/Last Update: 2014-05-04