Abstract

Extraction of high-quality proteins from tissue like Panax quinquefolius root is difficult due to high levels of interfering compounds, mainly viscous polysaccharide, which obstructs the extraction of low abundant proteins. To establish an efficient method for the application of proteomic analysis to P. quinquefolius root, 4 sample preparation methods were tested: Acetone precipitation, ethanol precipitation, chloroform/isoamyl alcohol extraction and phenol/chloroform/isoamyl alcohol extraction. These methods were evaluated based on the quality of SDS-PAGE and two-dimensional electrophoresis (2-DE) patterns. In chloroform/isoamyl alcohol extraction, due to the different polarity, proteins were dispersed in water-phase and inter-phase. There were abundant water-soluble proteins in water-phase and fat-soluble proteins in inter-phase. In order to acquire an optimized 2-DE pattern containing as many protein spots as it can be, the both phase proteins were mixed together, and the results of SDS-PAGE and 2-DE showed well-resolved low-abundant proteins with low background. Some significantly changed spots on 2-DE gel of optimized method were identified and their potential relationships to root development were discussed. These results showed that optimized method gives satisfactory and reproductive 2-DE protein patterns. It is expected that this method can also be applied to other plant root tissues that are rich in interfering ingredients.

Key words: 2-DE, MALDI-TOF/TOF MS, Panax quinquefolius root, protein extraction,proteomics.