Graduate Thesis Or Dissertation
 

Cytokinin production by Agrobacterium tumefaciens

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  • Certain plant-associated prokaryotes such as Corynebacteriurn fascians and Agrobacterium tumefaciens are known to produce substantial levels of cytokinins. In view of the fragmentary and presumptive evidence available on the quantitative and qualitative levels of cytokinins produced by A. tumefaciens, it was decided to conduct a study of the cytokinins produced by A. tumefaciens. In order to accomplish this objective, a sequence of methods was developed which allowed the isolation, purification and rigorous chemical identification of cytokinins at the sub-microgram level. The first step of purification utilized the adsorption of cytokinins onto C18 silica by a process known as trace enrichment. Further purification was effected by high performance liquid chromatography and cytokinin-active fractions were located by bioassay. Material in biologically active fractions was permethylated and analyzed by combined capillary gas-liquid chromatography and mass spectrometry. A. tumefaciens tRNA was found to contain . ⁶Ado, ms².⁶Ado, ms²io⁶Ado and trans-io⁶Ado. A. tumefaciens culture filtrates were found to contain i⁶Ade, ⁶Ado, ms²i ⁶Ado, ms²i o ⁶Ado and the trans isomers of io⁶ Ade and io ⁶Ado. Possible mechanisms for the production of these cytokinins were discussed. Preliminary evidence on three strains of A. tumefaciens showed that neither the production of N ⁶-Δ² -isopentenyladenine or zeatin correlated with the presence of the tumor-inducing plasmid.
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