塔玛亚历山大藻双向电泳蛋白的三种提取方法比较

Comparison of three protein extraction methods for proteomic analysis of Alexandrium tamarense with twodimensional electrophoresis

摘要:

摘要: 【目的】比较3 种蛋白质提取方法，找到适用于塔玛亚历山大藻蛋白的最佳的提取方法，为后续用双向电泳(2-DE) 技术研究不同条件下塔玛亚历山大藻蛋白的差异表达奠定基础。【方法】以塔玛亚历山大藻为研究对象，运用Tris-HCl 提取法、TCA 沉淀法和lysis buffer 提取法分别提取塔玛亚历山大藻蛋白，并通过双向电泳技术，对这3 种方法进行了比较分析，筛选出最适于塔玛亚历山大藻的蛋白提取方法。并运用以上得出的方法，以不加杀藻物质的无菌塔玛亚历山大藻为对照，比较分析了塔玛亚历山大藻在加入杀藻物质

Abstract:

Abstract: ［Objective］In order to find the best extraction method for proteins of Alexandrium tamarense for twodimensional electrophoresis (2-DE) analysis．［Methods］Three methods for extracting proteins from A．tamarense were compared，including trihydroxymethyl aminomethane (Tris-HCl) buffer extraction，trichloroacetic acid (TCA)/acetone precipitation and lysis buffer extraction． Alga was cultivated in normal f /2 media (control) and supplemented with algicidal substances． Proteins obtained using the best extraction method were separated with 2-DE． Protein-expression differences were identified using matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry (MS)．［Results］ Among the three protein extraction methods，lysis buffer extraction shows the best detection of the number and quality of protein spots with a clear background． Then，the lysis buffer extraction method was successfully applied to profiling protein expression in algicidal substances stress conditions and 14 differential expression proteins were identified using MALDI-TOF/MS．［Conclusion］Lysis buffer extraction was the most effective protein extraction method for Alexandrium tamarense．