Journal Article

Selective Excitation of Photosystems in Chloroplasts Inside Plant Leaves Observed by Near-Infrared Laser-Based Fluorescence Spectral Microscopy

Makoto Hasegawa, Takashi Shiina, Masahide Terazima and Shigeichi Kumazaki

in Plant and Cell Physiology

Published on behalf of Japanese Society of Plant Physiologists

Volume 51, issue 2, pages 225-238
Published in print February 2010 | ISSN: 0032-0781
Published online December 2009 | e-ISSN: 1471-9053 | DOI: http://dx.doi.org/10.1093/pcp/pcp182
Selective Excitation of Photosystems in Chloroplasts Inside Plant Leaves Observed by Near-Infrared Laser-Based Fluorescence Spectral Microscopy

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  • Biochemistry
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  • Plant Sciences and Forestry

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In this study, we produced selective images of photosystems in plant chloroplasts in situ. We used a spectroimaging microscope, equipped with a near-infrared (NIR) laser that provided light at wavelengths mainly between 800 and 830 nm, to analyze chlorophyll autofluorescence spectra and images from chloroplasts in leaves of Zea mays at room temperature. Femtosecond laser excitation of chloroplasts in mesophyll cells revealed a spectral shape that was attributable to PSII and its antenna in the centers of grana spots. We found that a continuous wave emitted by the NIR laser at a wavelength as long as 820 nm induced chlorophyll autofluorescence with a high contribution from PSI through a one-photon absorption mechanism. A spectral shape attributable to PSI and its antenna was thus obtained using continuous wave laser excitation of chloroplasts in bundle sheath cells. These highly pure spectra of photosystems were utilized for spectral decomposition at every intrachloroplast space to show distributions of PSI and PSII and their associated antenna. A new methodology using an NIR laser to detect the PSI/PSII ratio in single chloroplasts in leaves at room temperature is described.

Keywords: Chloroplast; Fluorescence spectra; Multiphoton microscopy; Photosystem; Thylakoid membrane; Zea mays

Journal Article.  9090 words.  Illustrated.

Subjects: Biochemistry ; Molecular and Cell Biology ; Plant Sciences and Forestry

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