The use of Trizol reagent (phenol/guanidine isothiocyanate) for producing high quality two-dimensional gel electrophoretograms (2-DE) of dinoflagellates.
Two-dimensional gel electrophoresis (2-DE) is one of the most efficient ways of resolving complex protein mixtures based on the isoelectric point (pI) and molecular mass (M(r)). Although it has been used extensively in proteomic studies on samples from the animal and plant kingdoms, there is limited information on its use on algae, such as dinoflagellates. The preparation of high-quality samples from dinoflagellate cells for 2-DE is difficult due to high endogenous levels of salts, nucleic acids, polysaccharides, phenolic compounds, pigments, and other interfering compounds. Desalting and concentrating steps are usually required for the preparation of dinoflagellate protein sample prior to 2-DE and these steps can be lengthy and complicated. In this study, we report the use of Trizol (a monophasic solution of phenol and guanidine isothiocyanate) for the extraction of proteins from dinoflagellate cells for 2-DE. The method is simple and fast. 2-DE profiles obtained with Trizol treatment are of very high quality in terms of resolution, spot number and spot intensity. This method greatly simplifies protein extraction procedures on dinoflagellate samples for obtaining a high quality and reproductive 2-DE profile. This methodology is generally applicable to algal samples.
Lee
Fred Wang-Fat
FW
The Proteomic Task Force, Department of Applied Biology and Chemical Technology, The Hong Kong Polytechnic University, Hung Hom, Hong Kong.
Lo
Samuel Chun-Lap
SC
D000096:Acetone / Q000032:analysis; D000818:Animals; D004141:Dinoflagellida / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D006146:Guanidines / Q000032:analysis*; D007202:Indicators and Reagents / Q000032:analysis; D017879:Isothiocyanates / Q000032:analysis*; D010449:Peptide Mapping; D019800:Phenol / Q000032:analysis*; D015800:Protozoan Proteins / Q000737:chemistry*; D020539:Sequence Analysis, Protein; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D006146:Guanidines; D007202:Indicators and Reagents; D017879:Isothiocyanates; D015800:Protozoan Proteins; C054435:guanidine isothiocyanate; D000096:Acetone; D019800:Phenol