An enzymatic fluorescent assay for the quantification of phosphite in a microtiter plate format

O. Berkowitz; R. Jost; S.J. Pearse; H. Lambers; P.M. Finnegan; G.E.St.J. Hardy; P.A. O’Brien

doi:10.1016/j.ab.2011.01.014

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An enzymatic fluorescent assay for the quantification of phosphite in a microtiter plate format

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An enzymatic fluorescent assay for the quantification of phosphite in a microtiter plate format

O. Berkowitz, R. Jost, S.J. Pearse, H. Lambers, P.M. Finnegan, G.E.St.J. Hardy and P.A. O’Brien

Analytical Biochemistry, Vol.412(1), pp.74-78

2011

DOI: https://doi.org/10.1016/j.ab.2011.01.014

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Abstract

A sensitive fluorometric assay for the quantification of phosphite has been developed. The assay uses the enzymatic oxidation of phosphite to phosphate by a recombinant phosphite dehydrogenase with NAD+ as cosubstrate to produce the highly fluorescent reaction product resorufin. The optimized assay can be carried out in a 96-well microtiter plate format for high-throughput screening purposes and has a detection limit of 0.25 nmol phosphite. We used the method to quantify phosphite levels in plant tissue extracts and to determine phosphite dehydrogenase activity in transgenic plants. The assay is suitable for other biological or environmental samples. Because phosphite is a widely used fungicide to protect plants from pathogenic oomycetes, the assay provides a cost-effective and easy-to-use method to monitor the fate of phosphite following application.

Details

Title: An enzymatic fluorescent assay for the quantification of phosphite in a microtiter plate format
Authors/Creators: O. Berkowitz (Author/Creator) - Murdoch University
R. Jost (Author/Creator) - The University of Western Australia
S.J. Pearse (Author/Creator) - The University of Western Australia
H. Lambers (Author/Creator) - The University of Western Australia
P.M. Finnegan (Author/Creator) - The University of Western Australia
G.E.St.J. Hardy (Author/Creator) - Murdoch University
P.A. O’Brien (Author/Creator) - Murdoch University
Publication Details: Analytical Biochemistry, Vol.412(1), pp.74-78
Publisher: Academic Press
Identifiers: 991005540099607891
Murdoch Affiliation: Centre for Phytophthora Science and Management
Language: English
Resource Type: Journal article

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Collaboration types: Domestic collaboration
Citation topics: 3 Agriculture, Environment & Ecology; 3.97 Plant Pathology; 3.97.1439 Phytophthora Management
Web Of Science research areas: Biochemical Research Methods; Biochemistry & Molecular Biology; Chemistry, Analytical
ESI research areas: Biology & Biochemistry