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Use of Fluorescent Reporters to Analyse Dynamic and Spatial Responses to Mechanical Wounding

  • Antoine Larrieu
  • Trang Hieu Nguyen
  • Antony ChampionEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 2085)

Abstract

Mechanical wounding of plant tissues triggers many different responses (Savatin DV, Gramegna G, Modesti V, Front Plant Sci 5:470, 2014). These are primarily mediated by the plant hormone Jasmonic Acid Isoleucine (JA-Ile). Recently, a fluorescent biosensor for JA-Ile showed that sample preparation (i.e., handling of samples) for fluorescent microscopy very often triggers wound response, even without apparent damage to the seedling, affecting downstream analyses (Larrieu A, Champion A, Legrand J, Nat Commun 6:6043, 2015). In this chapter, we describe how to overcome this technical limitation to monitor any fluorescent reporter or dye in response to wounding, using any type of fluorescent or confocal (inverted or upright, laser scanning or spinning disc) microscopes. Pharmacological or wound treatments can easily be performed and responses monitored over long periods of time. We further describe a simple method to extract and analyse quantitative data from confocal images using the open source software Fiji (Fiji Is Just ImageJ (Schindelin J, Arganda-Carreras I, Frise E, Nat Methods 9:676–682, 2012)) and OpenOffice.

Key words

Confocal microscopy Live imaging Jasmonic Acid Arabidopsis Wound responses Fluorescent biosensor 

Notes

Acknowledgments

A.L. is a postdoctoral fellow at the University of Leeds, UK. T.H.N. is supported by a French Embassy Fellowship.

References

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    Larrieu A, Champion A, Legrand J et al (2015) A fluorescent hormone biosensor reveals the dynamics of jasmonate signalling in plants. Nat Commun 6:6043CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2020

Authors and Affiliations

  • Antoine Larrieu
    • 1
  • Trang Hieu Nguyen
    • 2
  • Antony Champion
    • 3
    Email author
  1. 1.LS2 9JTLeedsUK
  2. 2.Institut de Recherche pour le Développement (IRD), CiradUniversité de Montpellier, UMR DIADEMontpellierFrance
  3. 3.UMR DIADE - IRDMontpellierFrance

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