Esterase isozyme studies in callus cultures

The enzyme polymorphism exhibited by isozyme patterns serves as a model for analyzing the gene action at different developmental stages, and the physiological status of the tissue differentiated to perform specified functions. Multiple molecular forms of a number of enzymes which were tissue specific were analyzed in inbred lines of maize (J. G. Scandalios, Proc. XII Int. Cong. Genet. 11:79-80, 1980). Tissue culture studies in our laboratory proved that the calli derived from immature embryos and glumes were capable of plantlet regeneration, whereas the seedling root calli failed to regenerate. The differences were mainly attributed to the physiological status and the extent of differentiation among the constituent cells of the explant cultures. Esterases exhibited polymorphism with different tissues at the whole plant level (Scandalios, J. Hered. 55:281, 1964). In the present study, regenerable calli derived from immature embryos and glumes and the nonregenerable calli of seedling root were compared for their esterase isozyme pattern.

Immature embryos (10-day-old) and glumes (60-day-old plants) of sweet corn were obtained from the field grown plants. Root explants were taken from one-week-old seedlings grown under aseptic conditions. Callus cultures were initiated on MS medium containing 2 mg/l 2,4-D. Four-week-old calli derived from the above explants were used for electrophoretic studies using standard PAGE technique. The gels were incubated for one hour in 100 ml of 0.6M phosphate buffer, pH 6.1, containing alpha-naphthyl acetate as substrate and fast blue RR as dye coupler to detect the esterases.

Significant differences were observed in esterase pattern in regenerable and non-regenerable calli. Calli of embryo, glume and root exhibited a total number of nine, eight and twelve bands, respectively, of which three common bands at Rf values 0.33, 0.46 and 0.70 were observed. The regenerable glume and embryo calli have shown one common band at Rf 0.22, which was absent in non-regenerable calli. The preliminary observations suggest that the presence of a specific band in regenerable, and its absence in nonregenerable calli, indicates a possible involvement of a specific esterase in differentiation.

P. S. Prasanna, K. V. Rao and G. M. Reddy

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