A TECHNIQUE FOR DETECTION OF CHITINASE, BETA-1,3-GLUCANASE, AND PROTEIN-PATTERNS AFTER A SINGLE SEPARATION USING POLYACRYLAMIDE-GEL ELECTROPHORESIS OR ISOELECTROFOCUSING

A procedure to detect chitinase and beta-1,3-glucanase isozymes and protein patterns after a single separation using native polyacrylamide gel electrophoresis (PAGE) or isoelectrofocusing (IEF) is described. After electrophoresis or isoelectrofocusing, an overlay gel containing glycol chitin as substrate for chitinase was incubated in close contact with the resolving gel. Chitinase isozymes were revealed by UV illumination after staining the overlay gel with fluorescent brightener 28. The resolving gel was then incubated with laminarin, and beta-1,3-glucanase isozymes were detected by using 2,3,5-triphenyltetrazolium chloride. The resolving gel with beta-1,3-glucanase bands was stained with Coomassie Brilliant Blue R 250 to reveal protein patterns. The isozymes were quantified by using native PAGE, and their pIs were estimated by IEF.