The most abundant protein (constituting ~47% of total soluble proteins) from the seeds of A. erioloba was a 53 kDa heat-stable protein, termed ASP53 (Fig. 6.1). Once purified to homogeneity it was used to immunize rabbits for antibody production. Immunocytochemistry using this antibody showed label to occur only in the cell wall of the axis (Fig. 6.2a) and the cell walls and protein storage vacuoles in cotyledons (Fig. 6.2b) of mature seeds. No gold particles, indicative of the presence of ASP53, were detected when similar sections were probed with the pre-immune serum (Fig. 6.2c). These locations suggest a dual role for ASP53, as a storage protein in the vacuoles and a possible stress-associated function in the cell wall.

Fig. 6.1. SDS-PAGE of total proteins extracted from mature seeds of Acacia erioloba (column 1), soluble proteins after incubation at 80°C for 30 min (column

2) and HPLC-purified protein (column

3). The molecular weights (in kDa) of markers (M) of known size are denoted on the left-hand side of the figure. The arrow shows the electrophoretic migration of the protein selected for further study.

to 31

to 31

1 2 Marker

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