Difference gel electrophoresis: a single gel method for detecting changes in protein extracts.
We describe a modification of two-dimensional (2-D) polyacrylamide gel electrophoresis that requires only a single gel to reproducibly detect differences between two protein samples. This was accomplished by fluorescently tagging the two samples with two different dyes, running them on the same 2-D gel, post-run fluorescence imaging of the gel into two images, and superimposing the images. The amine reactive dyes were designed to insure that proteins common to both samples have the same relative mobility regardless of the dye used to tag them. Thus, this technique, called difference gel electrophoresis (DIGE), circumvents the need to compare several 2-D gels. DIGE is reproducible, sensitive, and can detect an exogenous difference between two Drosophila embryo extracts at nanogram levels. Moreover, an inducible protein from E. coli was detected after 15 min of induction and identified using DIGE preparatively.
Unlü
M
M
Department of Chemistry, Center for Light Microscope Imaging and Biotechnology, Carnegie Mellon University, Pittsburgh, PA, USA.
Morgan
M E
ME
Minden
J S
JS
D000818:Animals; D004268:DNA-Binding Proteins; D004331:Drosophila melanogaster / Q000737:chemistry / Q000196:embryology; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D004926:Escherichia coli / Q000737:chemistry; D005047:Etoposide / Q000737:chemistry; D005456:Fluorescent Dyes; D011506:Proteins / Q000032:analysis*; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D029701:Saccharomyces cerevisiae Proteins; D017421:Sequence Analysis; D014157:Transcription Factors / Q000737:chemistry
D004268:DNA-Binding Proteins; D005456:Fluorescent Dyes; C052130:GAL4 protein, S cerevisiae; D011506:Proteins; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D005047:Etoposide