Effect of humic acid on callus culture of slash pine (Pinus elliottiiEngelm)1

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Abstract
An investigation was conducted to study the effect of humic acids (HA) on growth and nutrient uptake of slash pine (Pinus elliottii Engelm.) callus in vitro. Callus, differentiated from excised pine seed embryos, was grown at pH 5.7–5.8 for a total of nine weeks on a Brown and Lawrence culture medium to which 0 to 800 μg HA/mL were added. The treatments were replicated nine times. Samples of callus cultures were taken at periods of 3, 6 and 9 weeks, for size, fresh weight measurements, and macrostructure analyses by light microscopy, and scanning electron microscopy (SEM). Elemental composition of single callus cells was determined by x‐ray energy dispersive analysis (EDAX). SEM analysis indicated that HA had a slight effect on callus size. However, calli in 9 wk‐old cultures were significantly heavier than the control, whereas EDAX indicated that the P, K, Ca and S contents were increased considerably in individual callus cells over those in the control at 80, 160, and 200 μg HA/mL applications. The latter was attributed to the reaction of HA, increasing permeability of callus membranes, as reported for cellular plant membranes in the literature. The increased uptake, and consequently greater accumulation of nutrients in callus cells were the reasons for callus weight to increase in the presence of HA.