Modified TCA/acetone precipitation of plant proteins for proteomic analysis

Protein extracts obtained from cells or tissues often need to remove interfering substances for preparing high-quality protein samples in proteomic analysis. A number of protein extraction methods have been applied to various biological samples, especially TCA/acetone precipitation and phenol extraction. TCA/acetone precipitation, as a common method, is thought to minimize protein degradation and proteases activity as well as reduce contaminants like salts and polyphenols. However, the TCA/acetone precipitation relies on the completely pulverizing and repeatedly rinse of tissue powder to remove the interfering substances, which is laborious and time-consuming. In addition, a prolonged incubation in TCA/acetone or acetone can lead to the modifications and degradation of proteins, and the precipitated proteins are more difficult to re-dissolve. We have described a modified TCA/acetone precipitation of plant proteins for proteomic analysis. Proteins of cells or tissues were extracted using SDS-containing buffer, precipitated with equal volume of 20% TCA/acetone, and washed with acetone. Compared to classical TCA/acetone precipitation, this protocol generates comparable yields, spot numbers, and proteome profiling, but takes less time (ca. 45 min), thus avoiding excess protein modification and degradation after extended-period incubation in TCA/acetone or acetone. The modified TCA/acetone precipitation method is simple, fast, and suitable for various plant tissues in proteomic analysis.