Tracheid formation in cultures of pine (Pinus sylvestris)

Various combinations of growth factors were tested for their ability to induce phenylalanine ammonia lyase activity in a suspension culture of Pinus sylvestris L. Phenylalanine ammonia lyase is an established marker of differentiation in angiosperm tissue culture. A growth medium capable of inducing phenylalanine ammonia lyase activity was developed; it contained basal medium supplemented with 10mg 1⁻¹ (53.5 μM) naphthaleneacetic acid, 2mgl⁻¹ (9.3 μM) kinetin and 6% (175mM) sucrose. Using this medium it was possible to induce the differentiation of tracheids after 25 days in culture. Secondary thickening of the walls of cells cultured in the induction medium was confirmed by microscopical examination and correlated with an increase in the lignin content and changes in the polysacchar-ide composition of the cell wall. After culture in the induction medium there was an appreciable increase in the mannose content of wall polysaccharides, which suggested that synthesis of secondary wall glucomannan had been induced by the change in growth factor concentrations. High levels of xylose present in the cultured cells suggested the presence of a xyloglucan, which was subsequently isolated and characterized. ¹³C n.m.r. (nuclear magnetic resonance) spectroscopy was used to identify the glycosyl linkages in the xyloglucan.