Comparison of protein extraction methods suitable for proteomics analysis in seedling roots of Jerusalem artichoke under salt (NaCl) stress

Abstract

An efficient protein extraction method is a prerequisite for successful implementation of proteomics. In this study, seedling roots of Jerusalem artichoke were treated with the concentration of 250 mM NaCl for 36 h. Subsequently, six different protocols of protein extraction were applied to seedling roots of Jerusalem artichoke for comparing extraction efficiency by conducting two-dimensional electrophoresis (2-DE). The first-dimensional electrophoresis was differently performed by using tube gel and immobilized pH gradient (IPG) strips, respectively. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in the second dimension was equally carried out. The results indicated that precipitation of ammonium acetate in methanol after phenol extraction (Method 6) provided the best effect. Less impurity was contained in extracted proteins and the 2-DE gel images showed less horizontal and vertical stripes, the most proteineous spots were extracted on the basis of the protocol, up to 903. Trichloroacetic acid (TCA) method (Method 1) exhibited relative less quality gel images. 248 protein spots were only obtained according to the protocol and were the least among six protocols. Although, Mg/Nonidet P-40 (NP-40) methods (Methods 3 and 4) could extract a large number of proteins, it exhibited the worst quality gel images among all protocols and the horizontal and vertical stripes on the gel images were the most severe. It might be caused by a high salt content in extracted proteins.

Key words: Proteomics, Jerusalem artichoke, roots, two-dimensional electrophoresis (2-DE), protocol, salt.