Cytokinin analysis by immunoaffinity chromatography (IAC), high-performance liquid chromatography (HPLC) and radioimmunoassay (RIA) or enzyme-linked immunosorption assay (ELISA) has been used to study two separate topics: the role of tRNA in bacterial cytokinin biosynthesis and the changes in cytokinin concentration which occur during cereal grain development. Transfer RNA isopentenylation in the gall-forming plant pathogen Agrobacterium tumefaciens is encoded by the chromosomal miaA locus. Mutation of miaA reduces tRNA isopentenylation significantly and preliminary data suggest that turnover of isopentenylated tRNA is responsible for low level secretion of free N6-isopentenyladenine (iP) by the bacteria. However, the major route of cytokinin biosynthesis by gall-forming plant patho- genic bacteria is not via tRNA turnover but by direct biosynthesis mediated by dimethylallylpyro- phosphate: 5'-AMP transferase (DMAPP :AMP transferase) encoded by such genes as ipt, tzs (from A, tumefaciens) or ptz (from Pseudomonas savastanoi). Analysis of cytokinin levels in developing wheat and rice grains in the period immediately following pollination showed large transient increases in zeatin (Z) and zeatin riboside (ZR) which coincided with the period of maximum endosperm cell division reported by others. Detailed analyses of maize kernels, where development can be staged readily, showed that Z and ZR concentrations peaked 9 days after pollination (DAP). During the period 8-10 DAP, cytokinin oxidase underwent a significant increase in specific activity, indicating that cytokinin catabolism was enhanced as endosperm cell division ended.