Two-dimensional differential in-gel electrophoresis (DIGE) of leaf and roots of Lycopersicon esculentum.
In this report we present a detailed protocol for the analysis of differential protein expression between two plant tissue samples. The protocol involves harvesting of leaves and roots from mature tomato plants, preparing protein extracts from the harvested tissues, fluorescent labeling of each sample prior to differential in-gel electrophoresis (DIGE), first- and second-dimension electrophoretic separations, and image analysis to visualize and quantify differential protein expression. This protocol is adaptable for use with a wide variety of plant materials and can be used to measure protein expression changes occurring in response to abiotic stress, biotic stress, genetic manipulation, selective breeding, and many other conditions. In addition to the detailed protocol, we also present the results of a representative experiment analyzing subtle changes in protein expression in the roots of tomato plants grown under control and salt-stress conditions.
Keeler
Matthew
M
Department of Biochemistry and Molecular Biophysics, University of Arizona, Tucson, USA.
Letarte
Jessica
J
Hattrup
Emily
E
Hickman
Fatimah
F
Haynes
Paul A
PA
D000096:Acetone / Q000737:chemistry; D011232:Chemical Precipitation; D015180:Electrophoresis, Gel, Two-Dimensional / Q000295:instrumentation / Q000379:methods*; D004591:Electrophoresis, Polyacrylamide Gel; D005456:Fluorescent Dyes; D020869:Gene Expression Profiling / Q000379:methods; D007525:Isoelectric Focusing / Q000295:instrumentation / Q000379:methods; D018551:Solanum lycopersicum / Q000737:chemistry* / Q000378:metabolism; D018515:Plant Leaves / Q000737:chemistry / Q000378:metabolism; D010940:Plant Proteins / Q000737:chemistry / Q000302:isolation & purification*; D018517:Plant Roots / Q000737:chemistry / Q000378:metabolism; D040901:Proteomics / Q000295:instrumentation / Q000379:methods*; D012965:Sodium Chloride / Q000378:metabolism; D013194:Staining and Labeling; D014238:Trichloroacetic Acid / Q000737:chemistry
D005456:Fluorescent Dyes; D010940:Plant Proteins; D000096:Acetone; D012965:Sodium Chloride; D014238:Trichloroacetic Acid