Development of Pectic Projections on the Surface of Wound Callus Cells of Daucus carota L.

Abstract

During studies of wound healing in carrot roots, callus cells with prominent external projections were frequently observed. Protruberances of various sizes and shapes have previously been described on the outer wall surface of parenchyma cells in a number of vascular plants (Kisser, 1928; Carlquist, 1956), but were notably absent on wound callus cells of Daucus carota L. which developed on healed tissues of swollen tap roots (Sirtautaite and Sokolova, 1965) and carrot callus cells which had been grown axenically in various culture media (Fosket and Roberts, 1965; Israel and Steward, 1966; Halperin and Jensen, 1967). Carrots of the cultivars Bertina and Spalding Red Cored Improved were grown for 176 days under typical cultivation conditions in light peat soil, or in a coarse sand loam, as described previously (Davies and Lewis, 1980). Samples of 40 freshly-harvested roots of each cultivar were cut transversely into 5 cm thick discs and arranged in a single layer on nylon mesh over wet polystyrene foam in closed polythene boxes (80 x 45 x 8 cm) where the humidity, measured with lithium chloride sensors ('Hygrodynamics', V. A. Howe Ltd) was 0*95-0-98aw;. After periods of up to 10 days at 25 + 1 °C, sections of callus tissue 10-60 /?m thick were cut with a freezing microtome from the uppermost healed wound surface and examined with a Zeiss 'Universal' or 4R.A. 38' microscope with bright field, phase contrast and Nomarski interference systems. Histochemical tests with 0-5 per cent ammonium oxalate, zinc-chloro-iodine, Sudan IV, IKI-H2S04, Nile blue sulphate or osmium tetroxide were those described by Jensen (1962): hydroxylamine-ferric chloride by Reeve (1959); ruthenium red by Carlquist (1956); and methylene blue by Johansen (1940). For scanning electron microscopy (SEM) blocks (2x5 mm2) of callus tissue were carefully excised and mounted on aluminium stubs with silver 'Dag' (Acheson Colloid Company), and each stub was inserted into the surface of an aluminium block which had been precooled in liquid nitrogen. The rest of the procedure was as described by Lewis and Day (1972), and the material was examined with a 'Stereoscan Mk. II' (Cambridge Instruments Ltd) microscope. Specimens for transmission electron microscopy (TEM) were prepared by soaking the tissue either in unbuffered 2*5 per cent aqueous potassium permanaganate for 1 h, then washed in distilled water (Israel and Steward, 1966); or in 3 per cent gluteraldehyde containing 5 per cent sucrose, buffered with sodium cacodolyte at pH 7 0 (Jordan and Chapman, 1973), then in 2 per cent osmium tetroxide for 4 h. The material was then