| 引用本文: | 姜明国, 何小丹, 裴氏秋河, 江欢欢, 黄永颜, 陈保善.板栗疫病菌致病性机理的双向凝胶电泳法研究[J].广西植物,2008,(1):117-120.[点击复制] |
| JIANG Ming-Guo, HE Xiao-Dan, PEISHI Qiu-He, JIANG Huan-Huan,
HUANG Yong-Yan, CHEN Bao-Shan.Study on the pathogenicity of Cryphonectria parasitica by 2-DE[J].Guihaia,2008,(1):117-120.[点击复制] |
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| 板栗疫病菌致病性机理的双向凝胶电泳法研究 |
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姜明国1,4, 何小丹1, 裴氏秋河1, 江欢欢1, 黄永颜1, 陈保善1,2,3*
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1.广西大学 生命科学与技术学院, 南宁 530005;2.广西亚热带生物资源保护与利用重点实验室, 南宁 530005;3.微
生物及植物遗传工程教育部重点实验室, 南宁 530005;4.广西民族大学 化学与生态工程学院, 南宁 530007
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| 摘要: |
| 双向凝胶电泳技术是蛋白质组学研究的基础性技术平台。如何得到一张高质量的双向凝胶电泳图谱是进行后续研究的关键。为探索适用于板栗疫病菌可溶性总蛋白的最佳提取条件,从蛋白组学角度来探索板栗疫病菌致病性机理,比较了目前在丝状真菌中常用的两种蛋白质提取方法,制备的蛋白质样品经双向凝胶电泳后,在凝胶上呈现的蛋白质斑点的丰度和分布特点。结果表明,两种方法获得的蛋白质主要集中分布在pH4～7的范围内; TCA-丙酮沉淀法得到的图谱分辨率高但是蛋白质总量很少。裂解液-TCA-丙酮沉淀法得到的蛋白质总量较大,通过cleanup kit处理后图谱分辨率可以达到差异蛋白组的要求。随机提取几个银染蛋白点用MALDI-TOF MS/MS进行分析,可以得到高质量的肽质量指纹谱。表明该样品制备方法可以满足蛋白质鉴定的要求。 |
| 关键词: 蛋白质提取 双向凝胶电泳 蛋白质组 板栗疫病菌 |
| DOI： |
| 分类号:Q945.8 |
| 基金项目: |
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| Study on the pathogenicity of Cryphonectria parasitica by 2-DE |
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JIANG Ming-Guo1,4, HE Xiao-Dan1, PEISHI Qiu-He1, JIANG Huan-Huan1,
HUANG Yong-Yan1, CHEN Bao-Shan1,2,3*
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1.College of Life Science and Technology, China;2.Guangxi Key Laboratory of Subtropical Bioresource Conservation
and Utilization, China;3.Key Laboratory of Ministry of Education for Microbial and Plant Genetic Engineering,
Guangxi University, Nanning 530005, China;4.Guangxi University for nationality, Nanning 530007, China
1.College of Life Science and Technology, China; 2.Guangxi Key Laboratory of Subtropical Bioresource Conservation
and Utilization, China; 3.Key Laboratory of Ministry of Education for Microbial and Plant Genetic Engineering,
Guangxi University, Nanning 530005, China; 4.Guangxi University for nationality, Nanning 530007, China
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| Abstract: |
| Comprehensive protein extraction from Cryphonectria parasitica is a key step to achieve high quality protein separation in two dimensional electrophoresis(2-DE). Two routine cellular total protein extraction methods were compared in order to determine an optimal one for filamentous fungi. Then the extracted total proteins were subjected to 2-DE,and the better extraction method was determined by the indexes of protein distribution and abundance on corresponding silver-stained gel. Data showed that most proteins were distributed in the pH ranging from 4 to 7; Buffer-TCA-Acetone got much more whole protein,which was treated with cleanup kit could be separated very well by 2-DE.But TCA-Acetone method precipitated total protein which is less although which shows fine differentiation among protein spots. So we consider that Buffer-TCA-Acetone is appropriate protocol for C.parasitica. Several silver -stained protein spots were picked for MALDI-TOF MS/MS to identify and high-quality PMF and MS/MS spectras were obtained. Buffer-TCA-Acetone is suitable sample preparation protocol for C.parasitica proteomics. |
| Key words: protein extraction 2-DE proteomics Cryphonectria parasitica |
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