Biology Reference
In-Depth Information
Chapter 4
How to Use 2D Gel Electrophoresis in Plant Proteomics
Thierry Rabilloud
Abstract
Two-dimensional electrophoresis has nurtured the birth of proteomics. It is however no longer the exclusive
setup used in proteomics, with the development of shotgun proteomics techniques that appear more fancy
and fashionable nowadays.
Nevertheless, 2D gel-based proteomics still has valuable features, and sometimes unique ones, which
make it often an attractive choice when a proteomics strategy must be selected. These features are detailed
in this chapter, as is the rationale for selecting or not 2D gel-based proteomics as a proteomic strategy.
Key words
Two-dimensional electrophoresis, Proteomics, Blotting, Posttranslational modifi cations
1
Introduction
At the beginning of the twenty-fi rst century, it is fashionable to
describe 2D gel-based proteomics as an outdated and poorly effi -
cient technique. This feeling is mainly due to the fascination of
many scientists in proteomics for big lists, and in this case the well-
documented undersampling of 2D gel-based proteomics is dearly
resented, as well as its almost complete inability to analyze trans-
membrane proteins [
1
,
2
], despite of lot of effort devoted to this
particular issue (e.g. in [
3
,
4
] for plant samples). This undersam-
pling has been documented mostly on animal samples [
5
,
6
], but
it is obvious that it will take place on plant samples as well.
In fact, all proteomic techniques undersample complex sam-
ples by a ratio of 1:10 (1,000 proteins analyzed on 2D gels among
the 10,000 present in a cell, 20,000 peptides analyzed by shotgun
proteomics among the 200,000 derived from a cell sample).
However, as 2D gel-based proteomics is the only setup that ana-
lyzes complete proteins, the consequences of undersampling are
clear and heavy. If a protein is not seen, it is lost forever. In addi-
tion, the undersampling rules in 2D gel-based proteomics are well
known. Hydrophobic proteins, high molecular weight proteins,
and rare proteins do not show up.
