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Evaluation of extraction procedures for 2-DE analysis of aphid proteins

文献类型： 外文期刊

作者： Pan Yiou 1 ; An Shaoli 1 ; Li Kebin 2 ; Wang Tao 1 ; Fang Kui 1 ; Zhang Hua 1 ; Sun Yu 1 ; Yang Xun 1 ; Xi Jinghui 1 ;

作者机构： 1.Jilin Univ, Coll Plant Sci, Changchun 130062, Peoples R China

2.Chinese Acad Agr Sci, Inst Plant Protect, State Key Lab Biol Plant Dis & Insect Pests, Beijing 100193, Peoples R China

关键词： 2-DE;Protein extraction;Proteomics;Sitobion avenae

期刊名称：JOURNAL OF SEPARATION SCIENCE （ 影响因子：3.645； 五年影响因子：2.943 ）

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收录情况： SCI

摘要： Protein sample preparation is a crucial step in a 2-DE proteomics approach. In order to establish a routine protocol for the application of proteomics analysis to aphids, this study focuses on the specific protein extraction problems in insect tissues and evaluates four methods to bypass them. The approaches of phenol extraction methanol/ammonium acetate precipitation (PA), TCA/acetone precipitation, PEG precipitation, and no precipitation were evaluated for proteins isolation and purification from apterous adult aphids, Sitobion avenae. For 2-DE, the PA protocol was optimal, resulting in good IEF and clear spots. PA method yielded the greatest amount of protein and displayed most protein spots in 2-DE gels, as compared with the TCA/acetone precipitation, PEG precipitation and no precipitation protocols. Analysis of protein yield, image quality and spot numbers demonstrate that the TCA/acetone precipitation protocol is a reproducible and reliable method for extracting proteins from aphids. The PEG precipitation approach is a newly developed protein extraction protocol for aphids, from which more unique protein spots can be detected, especially for detection of acid proteins. These protocols are expected to be applicable to other insects or could be of interest to laboratories involved in insect proteomics, despite the amounts and types of interfering compounds vary considerably in different insects.

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