Graduate Thesis Or Dissertation
 

Substitution of germanium for boron in suspension-cultured carrot cells

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  • Boron has been recognized since 1923 as an essential micronutrient for vascular plants, but its function is still not known. Many diverse roles have been proposed, but none of these has been definitively shown. Problems that have made the study of the function of boron difficult include, no useful radioisotope, low level requirement (and toxicity at higher concentrations), and the wide spectrum of biochemical and physiological responses elicited. Germanium forms chemical complexes similar to those formed by boron, suggesting the possibility of using germanium as a tracer for boron. Previous studies (Mcllrath and Skok, 1966) with whole plants suggested that germanium could delay boron deficiency symptoms in sunflower seedlings for a few days, but not longer. To reevaluate this question, we have used suspension-cultured carrot cells and found that germanium can substitute for borate in this system. Germanate cultures were maintained by subculturing for more than two years. Cell growth was studied as a function of borate or germanate concentration of the medium. In addition, elemental analyses of B, Ge, Ca, Mg, Fe, Mn, Na, K, P, and Zn by Induction Coupled Plasma Argon Emission Spectroscopy were performed on these cells. To further characterize the role of boron, cells were fractionated into cell wall and protoplasmic fractions to determine the distribution of boron (or germanium) within the cell. Protoplasts were formed by enzymatic digestion of the cell wall. Alternatively, cells were fractionated by grinding in liquid nitrogen or passing through a French press, and centrifuging. These methods all yielded similar results, which showed that boron (or germanium) was concentrated in the cell wall fraction. At least 95% of the boron was present in the cell walls of cells grown in low boron concentrations. Germanium, was present in comparable concentrations in the cell wall, but was also found in significant amounts in the protoplasm. This suggests less than perfect control of the non-physiological element. One of the proposed roles for boron is as a structural member of the cell wall. The ability of germanium to substitute for boron in our system, and the similarity of chemical complexes formed by the two elements suggest a rather non-specific interaction with polyols. This, along with our finding that B (Ge) is localized in the cell wall are consistent with a role as a structural component in the cell wall.
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