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

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．

Key words:Keywords: Alexandrium tamarense，dinoflagellates，two-dimensional electrophoresis，protein extraction