Cytoskeletal dynamics in interphase, mitosis and cytokinesis analysed through Agrobacterium-mediated transient transformation of tobacco BY-2 cells
© 2010 The Authors. New Phytologist © 2010 New Phytologist Trust.
| Veröffentlicht in: | The New phytologist. - 1990. - 190(2011), 1 vom: 01. Apr., Seite 258-267 |
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| 1. Verfasser: | |
| Weitere Verfasser: | , , , |
| Format: | Online-Aufsatz |
| Sprache: | English |
| Veröffentlicht: |
2011
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| Zugriff auf das übergeordnete Werk: | The New phytologist |
| Schlagworte: | Journal Article Research Support, Non-U.S. Gov't Agrobacterium cell division cytoskeleton kinesin microtubule dynamics tobacco BY-2 transient gene expression Kinesins |
| Zusammenfassung: | © 2010 The Authors. New Phytologist © 2010 New Phytologist Trust. Transient transformation with Agrobacterium is a widespread tool allowing rapid expression analyses in plants. However, the available methods generate expression in interphase and do not allow the routine analysis of dividing cells. Here, we present a transient transformation method (termed 'TAMBY2') to enable cell biological studies in interphase and cell division. Agrobacterium-mediated transient gene expression in tobacco BY-2 was analysed by Western blotting and quantitative fluorescence microscopy. Time-lapse microscopy of cytoskeletal markers was employed to monitor cell division. Double-labelling in interphase and mitosis enabled localization studies. We found that the transient transformation efficiency was highest when BY-2/Agrobacterium co-cultivation was performed on solid medium. Transformants produced in this way divided at high frequency. We demonstrated the utility of the method by defining the behaviour of a previously uncharacterized microtubule motor, KinG, throughout the cell cycle. Our analyses demonstrated that TAMBY2 provides a flexible tool for the transient transformation of BY-2 with Agrobacterium. Fluorescence double-labelling showed that KinG localizes to microtubules and to F-actin. In interphase, KinG accumulates on microtubule lagging ends, suggesting a minus-end-directed function in vivo. Time-lapse studies of cell division showed that GFP-KinG strongly labels preprophase band and phragmoplast, but not the metaphase spindle |
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| Beschreibung: | Date Completed 31.12.2014 Date Revised 13.12.2023 published: Print-Electronic Citation Status MEDLINE |
| ISSN: | 1469-8137 |
| DOI: | 10.1111/j.1469-8137.2010.03587.x |