A quantitative fluorescence enzyme immunoassay for plant cytokinins.

An enzyme-linked immunosorbent assay (ELISA) which used 4-methylumbelliferyl phosphate as an enzyme substrate was used to quantify two plant cytokinins. This assay detected as little as 0.03 pmol (approximately 10 pg) of cytokinin in microplate wells coated with a cytokinin-ovalbumin conjugate. The method measured competition between free cytokinin and the bound conjugate for reaction with monoclonal anticytokinin antibodies and used a standard curve prepared by use of known amounts of free cytokinin to quantify hormone levels in unknown samples. Standard curves which consisted of logit/log plots of fluorescence units versus picomoles of competing cytokinin measured from 0.03 to 256 pmol (approximately 10-85,000 picograms) of zeatin riboside (ZR) or isopentenyl adenosine. The fluorescence ELISA was compared with radioimmunoassay for the quantification of ZR in wheat (Triticum aestivum L., cultivar Stephens) seed samples. This fluorescence ELISA method is recommended for use in combination with a fractionation method, such as HPLC, to quantify cytokinins present in plant extracts.

Metabolic changes associated with vernalization of wheat. I. Carbohydrate and nitrogen patterns.

A spring wheat (Triticum aestivum) and an obligate winter wheat (Triticum compactum) variety were each grown for 5 weeks in controlled environments at 2 degrees and 25 degrees . The threshold for flower induction in the winter wheat was 4 to 5 weeks at 2 degrees , whereas the spring wheat had no low temperature requirement for flowering. Changes in the levels of carbohydrate and nitrogen fractions in the wheat leaves were determined during their growth in the cold and warm environments. There was an enhanced accumulation of the 5 carbohydrate fractions in both wheat varieties grown at 2 degrees compared to 25 degrees . Highly significant differences in the levels of sucrose, oligosaccharides, and starch were found between the spring and winter varieties grown at 2 degrees . The winter wheat seedlings grown at 2 degrees accumulated much more of these carbohydrates than the corresponding spring wheat. The carbohydrate patterns in both varieties grown at 25 degrees were nearly identical except for the final 2 weeks of growth.The level of nitrogenous substances in the tissues grown at 2 degrees was much higher than in the corresponding tissues grown at 25 degrees . The only significant difference between the spring and winter varieties was in the soluble protein fraction. This fraction rose nearly 3-fold in the winter variety grown at 2 degrees , whereas it remained nearly constant in the similarly grown spring wheat. Most of the changing chemical patterns observed in relation to the vernalization treatment appear to be metabolic alterations associated with low temperature rather than alterations directly related with the vernalization response.