A New Bioassay for Auxins and Cytokinins

Abstract

The authors have developed a sensitive bioassay that can be used to detect auxins as well as cytokinins. The bioassay is based on the expression in transformed tobacco (Nicotiana tabacum) mesophyll protoplasts of a chimeric gene, consisting of the upstream sequences of the Agrobacterium tumefaciens gene 5, coupled to the coding sequence of the β-glucuronidase. The expression of this gene is induced by the presence of both auxin and cytokinin in the culture medium. Using this assay, indole-3-acetic acid was detected at 5 × 10−8 molar, whereas trans-zeatin could be detected at 5 × 10−11 molar. The assay can be performed in microtiter plates, allowing numerous samples to be analyzed simultaneously. Only 2.5 × 105 protoplasts are required for one individual assay in 250 microliters of culture medium and for qualitative results, the reaction is readily visualized by ultraviolet light.