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Imaging of viroids in nuclei from tomato leaf tissue by in situ hybridization and confocal laser scanning microscopy.

1989; Springer Nature; Volume: 8; Issue: 13 Linguagem: Inglês

10.1002/j.1460-2075.1989.tb08577.x

1460-2075

Jutta Harders, Nils Elias Lukacs, Michel Robert‐Nicoud, Thomas M. Jovin, Detlev Riesner,

Plant Pathogenic Bacteria Studies

Research Article20 December 1989free access Imaging of viroids in nuclei from tomato leaf tissue by in situ hybridization and confocal laser scanning microscopy. J. Harders J. Harders Heinrich-Heine-Universität Düsseldorf, Institut für Physikalische Biologie, FRG. Search for more papers by this author N. Lukács N. Lukács Heinrich-Heine-Universität Düsseldorf, Institut für Physikalische Biologie, FRG. Search for more papers by this author M. Robert-Nicoud M. Robert-Nicoud Heinrich-Heine-Universität Düsseldorf, Institut für Physikalische Biologie, FRG. Search for more papers by this author T.M. Jovin T.M. Jovin Heinrich-Heine-Universität Düsseldorf, Institut für Physikalische Biologie, FRG. Search for more papers by this author D. Riesner D. Riesner Heinrich-Heine-Universität Düsseldorf, Institut für Physikalische Biologie, FRG. Search for more papers by this author J. Harders J. Harders Heinrich-Heine-Universität Düsseldorf, Institut für Physikalische Biologie, FRG. Search for more papers by this author N. Lukács N. Lukács Heinrich-Heine-Universität Düsseldorf, Institut für Physikalische Biologie, FRG. Search for more papers by this author M. Robert-Nicoud M. Robert-Nicoud Heinrich-Heine-Universität Düsseldorf, Institut für Physikalische Biologie, FRG. Search for more papers by this author T.M. Jovin T.M. Jovin Heinrich-Heine-Universität Düsseldorf, Institut für Physikalische Biologie, FRG. Search for more papers by this author D. Riesner D. Riesner Heinrich-Heine-Universität Düsseldorf, Institut für Physikalische Biologie, FRG. Search for more papers by this author Author Information J. Harders1, N. Lukács1, M. Robert-Nicoud1, T.M. Jovin1 and D. Riesner1 1Heinrich-Heine-Universität Düsseldorf, Institut für Physikalische Biologie, FRG. The EMBO Journal (1989)8:3941-3949https://doi.org/10.1002/j.1460-2075.1989.tb08577.x PDFDownload PDF of article text and main figures. ToolsAdd to favoritesDownload CitationsTrack CitationsPermissions ShareFacebookTwitterLinked InMendeleyWechatReddit Figures & Info The intracellular localization of viroids has been investigated by viroid-specific in situ hybridization and analysis by digital microscopy of the distribution of the fluorescent hybridization signals. Isolated nuclei from green leaf tissue of tomato plants infected with potato spindle tuber viroid (PSTVd) were bound to microscope slides, fixed with formaldehyde and hybridized with biotinylated transcripts of cloned PSTVd cDNA. The bound probe was detected with lissamine--rhodamine conjugated streptavidin. Nucleoli were identified by immunofluorescence using the monoclonal antibody Bv96 and a secondary FITC-conjugated antibody. In plants infected with either a lethal or an intermediate PSTVd strain, the highest intensity of fluorescence that arose from hybridization with the probe specific for the viroid (+)strand was found in the nucleoli, confirming results of previous fractionation studies. A similar distribution was found for (-)strand replication intermediates of PSTVd using specific (+)strand transcripts as hybridization probes. In order to determine if viroids are located at the surface or in the interior of the nucleoli, the distribution of the fluorescence hybridization signals was studied with a confocal laser scanning microscope (CLSM). It was shown by three-dimensional reconstruction that viroids are neither restricted to the surface of the nucleoli nor to a peripheral zone, but are instead homogeneously distributed throughout the nucleolus. The functional implications of the intranucleolar location of viroids and their replication intermediates are discussed with respect to proposed mechanisms of viroid replication and pathogenesis. Previous ArticleNext Article Volume 8Issue 131 December 1989In this issue RelatedDetailsLoading ...