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JXB Advance Access published online on April 4, 2006
Journal of Experimental Botany, doi:10.1093/jxb/erj181
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© The Author [2006]. Published by Oxford University Press [on behalf of the Society for Experimental Biology]. All rights reserved. For Permissions, please e-mail: journals.permissions@oxfordjournals.org
Received December 15, 2005
Accepted March 3, 2006

Oxygen Metabolism Special Issue Article

Imaging the production of singlet oxygen in vivo using a new fluorescent sensor, Singlet Oxygen Sensor Green®

Cristina Flors 1, Michael J. Fryer 2, Jen Waring 2, Brandon Reeder 2, Ulrike Bechtold 2, Philip M. Mullineaux 2, Santi Nonell 3, Michael T. Wilson 2, and Neil R. Baker 2 *

1 Department of Biological Sciences, University of Essex, Colchester, Essex CO4 3SQ, UK; Grup d'Enginyeria Molecular, Institut Químic de Sarria, Universitat Ramon Lull, Via Augusta 390, E-08017 Barcelona, Spain; Present address: Department of Chemistry, Katholieke Universiteit Leuven, Celestijnenlaan 200F, B-3001 Heverlee, Belgium
2 Department of Biological Sciences, University of Essex, Colchester, Essex CO4 3SQ, UK
3 Grup d'Enginyeria Molecular, Institut Químic de Sarria, Universitat Ramon Lull, Via Augusta 390, E-08017 Barcelona, Spain

* To whom correspondence should be addressed.
Neil R. Baker, E-mail: baken{at}essex.ac.uk


  Abstract

Singlet oxygen is known to be produced by cells in response to photo-oxidative stresses and wounding. Due to singlet oxygen being highly reactive, it is thought to have a very short half-life in biological systems and, consequently, it is difficult to detect. A new commercially available reagent (singlet oxygen sensor green, SOSG), which is highly selective for singlet oxygen, was applied to a range of biological systems that are known to generate singlet oxygen. Induction of singlet oxygen production by the addition of myoglobin to liposome preparations demonstrated that the singlet oxygen-induced increases in SOSG fluorescence closely followed the increase in the concentration of conjugated dienes, which is stoichiometrically related to singlet oxygen production. Applications of photo-oxidative stresses to diatom species and leaves, which are known to result in the production of singlet oxygen, produced large increases in SOSG fluorescence, as did the addition of 3-(3',4'-dichlorophenyl)1,1-dimethylurea (DCMU) to these systems, which inhibits electron transport in photosystem II and stimulates singlet oxygen production. The conditional fluorescent (flu) mutant of Arabidopsis produces singlet oxygen when exposed to light after a dark period, and this coincided with a large increase in SOSG fluorescence. Wounding of leaves was followed by an increase in SOSG fluorescence, even in the dark. It is concluded that SOSG is a useful in vivo probe for the detection of singlet oxygen.

Keywords: Arabidopsis; diatoms; flu mutant; herbicides; liposomes; photo-oxidative stress; singlet oxygen; wounding.
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