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Effective protein extraction protocol for proteomics studies of Jerusalem artichoke leaves

文献类型： 外文期刊

作者： Zhang, Meide 2 ; Shen, Shihua 1 ;

作者机构： 1.Chinese Acad Sci, Inst Bot, Beijing, Peoples R China

2.Hubei Acad Agr Sci, Inst Chinese Herbal Med, Enshi 44500, Peoples R China

关键词： 2DE;Jerusalem artichoke;Protein extraction;Proteomics

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 extraction is a crucial step for proteomics studies. To establish an effective protein extraction protocol suitable for two-dimensional electrophoresis (2DE) analysis in Jerusalem artichoke (Helianthus tuberosus L.), three different protein extraction methods - trichloroacetic acid/acetone, Mg/NP-40, and phenol/ammonium acetate - were evaluated using Jerusalem artichoke leaves as source materials. Of the three methods, trichloroacetic acid/acetone yielded the best protein separation pattern and highest number of protein spots in 2DE analysis. Proteins highly abundant in leaves, such as Rubisco, are typically problematic during leaf 2DE analysis, however, and this disadvantage was evident using trichloroacetic acid/acetone. To reduce the influence of abundant proteins on the detection of low-abundance proteins, we optimized the trichloroacetic acid/acetone method by incorporating a PEG fractionation approach. After optimization, 363 additional (36.2%) protein spots were detected on the 2DE gel. Our results suggest that trichloroacetic acid/acetone method is a better protein extraction technique than Mg/NP-40 and phenol/ammonium acetate in Jerusalem artichoke leaf 2DE analysis, and that trichloroacetic acid/acetone method combined with PEG fractionation procedure is the most effective approach for leaf 2DE analysis of Jerusalem artichoke.

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