Direct detection of beta-1,3-glucanase isozymes on polyacrylamide electrophoresis and isoelectrofocusing gels.
A procedure to assay isozymes of beta-1,3-glucanase directly on polyacrylamide gel electrophoresis (PAGE) and isoelectrofocusing (IEF) gels by using 2,3,5-triphenyltetrazolium chloride is described. The reagent reacts with reducing sugars released by beta-1,3-glucanases from the substrate laminarin. Acidic and neutral isozymes of beta-1,3-glucanase were detected and quantified on 17.5% native PAGE gels run with an anodic buffer system. A significant linear relationship (alpha = less than 0.01, R = 0.991) was observed between amounts of beta-1,3-glucanase loaded and intensity of bands stained with the reagent on native PAGE gels. A full isozyme pattern was obtained on 7.5% IEF gels with a pH range of 3.5-9.5. The IEF gels were heated in a microwave oven during the staining process to minimize diffusion.
Pan
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Department of Plant Pathology, University of Kentucky, Lexington 40546.
Ye
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Kuć
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D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D043326:Glucan 1,3-beta-Glucosidase; D005959:Glucosidases / Q000032:analysis*; D007525:Isoelectric Focusing / Q000379:methods; D007527:Isoenzymes / Q000032:analysis*; D013194:Staining and Labeling; D013778:Tetrazolium Salts; D001617:beta-Glucosidase / Q000032:analysis*
D007527:Isoenzymes; D013778:Tetrazolium Salts; C009591:triphenyltetrazolium; D005959:Glucosidases; D001617:beta-Glucosidase; D043326:Glucan 1,3-beta-Glucosidase