Plant tissues contain large amounts of secondary compounds that significantly interfere with
protein extraction and 2DE analysis. Thus, sample preparation is a crucial step prior to 2DE in
plant proteomics. This tutorial highlights the guidelines that need to be followed to performan
adequate total proteinextractionbefore 2DE inplant proteomics.Webrieflydescribe thehistory,
development, and feature of major sample preparation methods for the 2DE analysis of plant
tissues, that is, trichloroacetic acid/acetone precipitation and phenol extraction. We introduce
the interfering compounds in plant tissues and the general guidelines for tissue disruption,
protein precipitation and resolubilization. We describe in details the advantages, limitations,
and application of the trichloroacetic acid/acetone precipitation and phenol extraction methods
to enable the readers to select the appropriate method for a specific species, tissue, or cell
type. The current applications of the sample preparation methods in plant proteomics in the
literature are analyzed. A comparative proteomic analysis between male and female plants
ofPistacia chinensisis used as an example to represent the sample preparation methodology
in 2DE-based proteomics. Finally, the current limitations and future development of these
sample preparationmethods are discussed. This Tutorial is part of the International Proteomics
Tutorial Programme (IPTP17).