Abstract
Laser capture microdissection (LCM) has become a powerful technique that allows analyzing gene expression in specific target cells from complex tissues. It is widely used in animal research, yet few studies on plants have been carried out. We have applied this technique to the plants–nematode interaction by isolating feeding cells (giant cells; GCs) immersed inside complex swelled root structures (galls) induced by root-knot nematodes. For this purpose, a protocol that combines good morphology preservation with RNA integrity maintenance was developed, and successfully applied to Arabidopsis and tomato galls. Specifically, early developing GCs at 3 and 7 days post infection (dpi) were analyzed; RNA from LCM GCs was amplified and used successfully for microarray assays.
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Acknowledgments
The authors thank K. Lindsey and J. Topping (University of Durham, UK) for the use of their equipment and for their advice during the development of the microdissection protocol. Work in the laboratory was supported by grants from the Ministry of Education (AGL2007-60273) and the Ministry of Science and Innovation (AGL2010-17388) to CE, and PCI08-0074-0294 from the Junta de Comunidades de Castilla la Mancha and CSD2007-00057 from the Ministry of Science and Innovation to CF. M.B. was a recipient of a postdoc fellowship from the ConsejerĂa of Science and Education (JCMM) and the European Social Fund.
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Barcala, M., Fenoll, C., Escobar, C. (2012). Laser Microdissection of Cells and Isolation of High-Quality RNA After Cryosectioning. In: Jin, H., Gassmann, W. (eds) RNA Abundance Analysis. Methods in Molecular Biology, vol 883. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-61779-839-9_6
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DOI: https://doi.org/10.1007/978-1-61779-839-9_6
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Online ISBN: 978-1-61779-839-9
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