Abstract
Here, we describe a new imaging setup able to assess in vivo photosynthetic activity. The system specifically measures time-resolved chlorophyll fluorescence in response to light. It is composed of a fast digital camera equipped with a wide-angle lens for the analysis of samples up to 10 × 10 cm, i.e. entire plants or petri dishes. In the choice of CCD, we have opted for a 12-bits high frame rate [150 fps (frames per second)] at the expense of definition (640 × 480 pixels). Although the choice of digital camera is always a compromise between these two related features, we have designed a flexible system allowing the fast sampling of images (down to 100 μs) with a maximum spatial resolution. This image readout system, synchronized with actinic light and saturating pulses, allows a precise determination of F 0 and F M, which is required to monitor PSII activity. This new imaging system, together with image processing techniques, is useful to investigate the heterogeneity of photosynthetic activity within leaves or to screen large numbers of unicellular algal mutant colonies to identify those with subtle changes in photosynthetic electron flow.
Abbreviations
- CCD:
-
Charge-coupled device
- Pixel:
-
Picture element
- LED:
-
Light-emitting diode
- cyt:
-
Cytochrome
- PSI:
-
Photosystem I
- PSII:
-
Photosystem II
- PQ:
-
Plastoquinone
- PQH2 :
-
Plastoquinol
References
Baker NR (2008) Chlorophyll fluorescence: a probe of photosynthesis in vivo. Annu Rev Plant Biol 59:89–113
Bennoun P, Béal D (1997) Screening algal mutant colonies with altered thylakoid electrochemical gradient through fluorescence and delayed luminescence digital imaging. Photosynth Res 51(2):161–165
Bennoun P, Béal D (1998). New digital imaging instrument for measuring fluorescence and delayed luminescence. In: Rochaix J-D, Goldschmidt-Clermont M, Merchant S (eds) The molecular biology of chloroplasts and mitochondria in Chlamydomonas, vol 7. Advances in photosynthesis. Kluwer Academic Publishers, Dordrecht, The Netherlands, pp 451–458
Chen X, Kindle K et al (1993) Initiation codon mutations in the Chlamydomonas chloroplast petD gene result in temperature-sensitive photosynthetic growth. EMBO J 12(9):3627–3635
Cuni A, Xiong L et al (2004) Modification of the pheophytin midpoint potential in photosystem II: modulation of the quantum yield of charge separation and of charge recombination pathways. Phys Chem Chem Phys 6:4825–4831
de Vitry C, Desbois A et al (2004) Biochemical and spectroscopic characterization of the covalent binding of heme to cytochrome b 6. Biochemistry 43(13):3956–3968
Joliot P, Béal D et al (1980) Une nouvelle méthode spectrophotométrique destinée à l’étude des réactions photosynthétiques. J Chim Phys 77(3):209–216
Kuras R, Wollman FA (1994) The assembly of cytochrome b6/f complexes: an approach using genetic transformation of the green alga Chlamydomonas reinhardtii. EMBO J 13(5):1019–1027
Nedbal L, Soukupova J et al (2000) Kinetic imaging of chlorophyll fluorescence using modulated light. Photosynth Res 66(1–2):3–12
Saint-Marcoux D, Wollman F-A et al. (2009) Biogenesis of cytochrome b 6 in photosynthetic membranes. J Cell Biol 85(7):1195–1207
Schreiber U, Quayle P et al (2007) Methodology and evaluation of a highly sensitive algae toxicity test based on multiwell chlorophyll fluorescence imaging. Biosens Bioelectron 22(11):2554–2563
Acknowledgements
The authors are indebted to Catherine de Vitry for the SUIV and CF0ß antibodies and to David Stern for the kind gift of iniD mutants. Financial support is acknowledged from Fondation Pierre-Gilles de Gennes pour la recherche and Agence Nationale pour la Recherche ANR-08-BIOE-002 ALGOMICS.
Author information
Authors and Affiliations
Corresponding author
Electronic supplementary material
Below is the link to the electronic supplementary material.
Supplementary material [The animated version of Figure 3B is available as a movie (mpeg 1 format).] (MPEG 1684 kb)
Rights and permissions
About this article
Cite this article
Johnson, X., Vandystadt, G., Bujaldon, S. et al. A new setup for in vivo fluorescence imaging of photosynthetic activity. Photosynth Res 102, 85–93 (2009). https://doi.org/10.1007/s11120-009-9487-2
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s11120-009-9487-2