Two-dimensional electrophoresis of acidic proteins isolated from ozone-stressed Norway spruce needles (Picea abies L. Karst): separation method and image processing.
The effect of ozone stress (< 10,200 or 400 micrograms m-3) on the protein pattern of Picea abies L. Karst needles was studied by fumigating potted grafts from mature trees. Clonal trees of Norway spruce were fumigated in open-top chambers for one growing season. Proteins, soluble and membrane bound, were extracted from the needles in a four-step procedure: (i) grinding in liquid nitrogen and then in a pH 5.0 buffer with thiourea added to inhibit phenol oxidase and polyvinylpolypyrrolidone (PVP) added to bind phenolic compounds; (ii) precipitation of proteins with acetone at -20 degrees C; (iii) solubilization in a modified lysis buffer; and (iv) dialysis against the modified lysis buffer. Proteins were separated by two-dimensional electrophoresis according to O'Farrell's method. The changes in the silver-stained protein pattern were evaluated by using image processing methods. Electrophoretograms from stressed and nonstressed trees are presented.
D002264:Carboxylic Acids / Q000302:isolation & purification*; D015180:Electrophoresis, Gel, Two-Dimensional; D005069:Evaluation Studies as Topic; D005651:Fumigation; D007091:Image Processing, Computer-Assisted; D007525:Isoelectric Focusing; D010126:Ozone / Q000633:toxicity*; D010940:Plant Proteins / Q000302:isolation & purification*; D012016:Reference Values; D016622:Silver Staining; D014197:Trees
D002264:Carboxylic Acids; D010940:Plant Proteins; D010126:Ozone