The Xylem Ureide Assay of Nitrogen Fixation: Sampling Procedures and Sources of Error

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Abstract
Herridge, D. F., O'Connell, P. and Donnelly, K. 1988. The xylem ureide assay of nitrogen fixation: sampling procedures and sources of error.—J. exp. BoL 39: 12–22. The relative abundance of ureides [(ureide-N/ureide-N + NO3–−N + α amino-N)× 100] in vacuum-extracted xylem exudate of soyabean (Glycine max [L.] Merr.) can be used to quantify N2 fixation activity. We report experiments designed to (1) develop appropriate procedures for sampling and storage of xylem exudate extracted from plants in the field and (2) develop a single plant, non-destructive sampling procedure, useful for evaluation of segregating breeding material. Soyabean cultivars Bragg, Essex, Davis, Hardee and Dodds were grown at three sites at Brecza, New South Wales, on a deep, alkaline vertisol. All plots were inoculated with effective Bradyrhizobium japonicum CB 1809. Vacuum strengths of between 50⋅5 and 80⋅8 kN m−2 did not affect the relative abundance of ureides of xylem exudate. Delays between plant sampling and extraction of exudate, however, resulted in time-dependent increases in the relative ureide contents. Extraction should immediately follow shoot detachment to allow valid comparison of treatments and accurate interpretation of data. The relative ureide contents of exudate showed only minor diurnal fluctuations. Exudate, once collected, could be stabilized by dilution (1:1) in absolute ethanoL Both diluted and undiluted samples were stable for at least 7 d at 4°C. Differences during early flowering, but not subsequently, in the compositions of N solutes of exudate extracted from either the whole shoot, the top half or the lower half indicated the possibility of non-destructive sampling where seed, as well as xylem exudate, could be harvested. Best results in terms of volume of exudate extracted, treatment separation and seed production were obtained when single plants were sampled during late flowering by detachment of shoots at internode 5.