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Plant Developmental Biology pp 321–346Cite as

Laser Microdissection of Paraffin-Embedded Plant Tissues for Transcript Profiling

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 655))

Abstract

High-resolution cellular analysis will help answer many important questions in plant biology including how genetic information is differentially used to enable the formation and development of the plant body. By comparing transcriptome data from distinct cell types during various stages of development, insight can be obtained into the transcriptional networks that underpin the attributes and contributions of particular cells and tissues. Laser microdissection (LM) is a technique that enables researchers to obtain specific cells or tissues from histological samples in a manner conducive to downstream molecular analysis. LM has become an established strategy in many areas of biology and it has recently been adapted for use with many types of plant tissue.

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References

  1. Schmid, M., Davison, T. S., Henz, S. R., et al. (2005) A gene expression map of Arabidopsis thaliana development. Nat Gen 37, 501–506.

    Article  CAS  Google Scholar 

  2. Benedito, V. A., Torres-Jerez, I., Murray, J. D., et al. (2008) A gene expression atlas of the model legume Medicago truncatula. Plant J 55, 504–513.

    Article  PubMed  CAS  Google Scholar 

  3. Jiao, Y., Lori Tausta, S., Gandotra, N., et al. (2009) A transcriptome atlas of rice cell types uncovers cellular, functional and developmental hierarchies. Nat Gen 41, 258–263.

    Article  CAS  Google Scholar 

  4. Balestrini, R. and Bonfante, P. (2008) Laser microdissection (LM): Applications to plant materials. Plant Biosyst 142, 331–336.

    Article  Google Scholar 

  5. Day, R. C., Grossniklaus, U., and Macknight, R. C. (2005) Be more specific! Laser-assisted microdissection of plant cells. Trends Plant Sci 10, 397–406.

    Article  PubMed  CAS  Google Scholar 

  6. Nelson, T., Tausta, S. L., Gandotra, N., and Liu T. (2006) Laser microdissection of plant tissue: What you see is what you get. Annu Rev Plant Biol 57, 181–201.

    Article  PubMed  CAS  Google Scholar 

  7. Ohtsu, K., Takahashi, H., Schnable, P., and Nakazono, M. (2006) Cell Type-specific gene expression profiling in plants by using a combination of laser microdissection and high-throughput technologies. Plant Cell Physiol 48, 3–7.

    Article  PubMed  Google Scholar 

  8. Cai, S. and Lashbrook, C. (2008) Stamen abscission zone transcriptome profiling reveals new candidates for abscission control: enhanced retention of floral organs in transgenic plants overexpressing Arabidopsis ZINC FINGER PROTEIN2. Plant Physiol 146, 1305–1321.

    Article  PubMed  CAS  Google Scholar 

  9. Day, R. C., Herridge, R. P., Ambrose, B. A., and Macknight, R. C. (2008) Transcriptome analysis of proliferating Arabidopsis endosperm reveals biological implications for the control of syncytial division, cytokinin signaling, and gene expression regulation. Plant Physiol 148, 1964–1984.

    Article  PubMed  CAS  Google Scholar 

  10. Ohtsu, K., Smith, M., Emrich, S., et al. (2007) Global gene expression analysis of the shoot apical meristem of maize (Zea mays L.). Plant J 52, 391–404.

    Article  PubMed  CAS  Google Scholar 

  11. Wu, Y., Machado, A. C., White, R. G., Llewellyn, D., and Dennis, E. (2006) Expression profiling identifies genes expressed early during lint fibre initiation in cotton. Plant Cell Physiol 47, 107–127.

    Article  PubMed  CAS  Google Scholar 

  12. Galbraith, D. W. (2003) Global analysis of cell type-specific gene expression. Comp Funct Genom 4, 208–215.

    Article  CAS  Google Scholar 

  13. Nelson, T., Gandotra, N., and Tausta, S. (2008) Plant cell types: Reporting and sampling with new technologies. Curr Opin Plant Biol 11, 567–573.

    Article  PubMed  CAS  Google Scholar 

  14. Day, R., Mcnoe, L., and Macknight, R. (2007) Transcript analysis of laser microdissected plant cells. Physiol Plant 129, 267–282.

    Article  CAS  Google Scholar 

  15. Espina, V., Wulfkuhle, J. D., Calvert, V. S., et al. (2006) Laser-capture microdissection. Nat Protoc 1, 586–603.

    Article  PubMed  CAS  Google Scholar 

  16. Ivashikina, N., Deeken, R., Ache, P., et al. (2003) Isolation of AtSUC2 promoter-GFP-marked companion cells for patch-clamp studies and expression profiling. Plant J 36, 931–945.

    Article  PubMed  CAS  Google Scholar 

  17. Casson, S., Spencer, M., Walker, K., and Lindsey, K. (2005) Laser capture microdissection for the analysis of gene expression during embryogenesis of Arabidopsis. Plant J 42, 111–123.

    Article  PubMed  CAS  Google Scholar 

  18. Spencer, M. W., Casson, S. A., and Lindsey, K. (2007) Transcriptional profiling of the Arabidopsis embryo. Plant Physiol 143, 924–940.

    Article  PubMed  CAS  Google Scholar 

  19. Inada, N. and Wildermuth, M. C. (2005) Novel tissue preparation method and cell-specific marker for laser microdissection of Arabidopsis mature leaf. Planta 221, 9–16.

    Article  PubMed  CAS  Google Scholar 

  20. Cai, S. and Lashbrook, C. C. (2006) Laser capture microdissection of plant cells from tape-transferred paraffin sections promotes recovery of structurally intact RNA for global gene profiling. Plant J 48, 628–637.

    Article  PubMed  CAS  Google Scholar 

  21. Deeken, R., Ache, P., Kajahn, I., Klinkenberg, J., Bringmann, G., and Hedrich, R. (2008) Identification of Arabidopsis thaliana phloem RNAs provides a search criterion for phloem-based transcripts hidden in complex datasets of microarray experiments. Plant J 55, 746–759.

    Article  PubMed  CAS  Google Scholar 

  22. Day, R. C., McNoe, L., and Macknight, R. C. (2007) Evaluation of global RNA amplification and its use for high-throughput transcript analysis of laser-microdissected endosperm. Int J Plant Genom 61028.

    Google Scholar 

  23. Galbiati, M., Simoni, L., Pavesi, G., et al. (2008) Gene trap lines identify Arabidopsis genes expressed in stomatal guard cells. Plant J 53, 750–762.

    Article  PubMed  CAS  Google Scholar 

  24. Thiel, J., Weier, D., Sreenivasulu, N., et al. (2008) Different hormonal regulation of cellular differentiation and function in nucellar projection and endosperm transfer cells: a microdissection-based transcriptome study of young barley grains. Plant Physiol 148, 1436–1452.

    Article  PubMed  CAS  Google Scholar 

  25. Wu, Y., Llewellyn, D., White, R., Ruggiero, K., Al-Ghazi, Y., and Dennis, E. (2007) Laser capture microdissection and cDNA microarrays used to generate gene expression profiles of the rapidly expanding fibre initial cells on the surface of cotton ovules. Planta 226, 1475–1490.

    Article  PubMed  CAS  Google Scholar 

  26. Santi, S. and Schmidt, W. (2008) Laser microdissection-assisted analysis of the functional fate of iron deficiency-induced root hairs in cucumber. J Exp Bot 59, 697–704.

    Article  PubMed  CAS  Google Scholar 

  27. Jiang, K., Zhang, S., Lee, S., et al. (2006) Transcription profile analyses identify genes and pathways central to root cap functions in maize. Plant Mol Biol 60, 343–363.

    Article  PubMed  CAS  Google Scholar 

  28. Dembinsky, D., Woll, K., Saleem, M., et al. (2007) Transcriptomic and proteomic analyses of pericycle cells of the maize primary root. Plant Physiol 145, 575–588.

    Article  PubMed  CAS  Google Scholar 

  29. Nakazono, M., Qiu, F., Borsuk, L. A., et al. (2003) Laser-capture microdissection, a tool for the global analysis of gene expression in specific plant cell types: Identification of genes expressed differentially in epidermal cells or vascular tissues of maize. Plant Cell 15, 583–596.

    Article  PubMed  CAS  Google Scholar 

  30. Zhang, X., Madi, S., Borsuk, L., et al. (2007) Laser microdissection of narrow sheath mutant maize uncovers novel gene expression in the shoot apical meristem. PLoS Genet 3, e101.

    Article  PubMed  Google Scholar 

  31. Nakada, M., Komatsu, M., Ochiai, T., et al. (2006) Isolation of from and analysis of its expression using laser microdissection. Plant Sci 170, 143–150.

    Article  CAS  Google Scholar 

  32. Corpas, F. J., Fernandez-Ocana, A., Carreras, A., et al. (2006) The expression of different superoxide dismutase forms is cell-type dependent in olive (Olea europaea L.) leaves. Plant Cell Physiol 47, 984–994.

    Article  PubMed  CAS  Google Scholar 

  33. Murata, J. and Luca, V. (2005) Localization of tabersonine 16-hydroxylase and 16-OH tabersonine-16-O-methyltransferase to leaf epidermal cells defines them as a major site of precursor biosynthesis in the vindoline pathway in Catharanthus roseus. Plant J 44, 581–594.

    Article  PubMed  CAS  Google Scholar 

  34. Yu, Y., Lashbrook, C. C., and Hannapel, D. J. (2007) Tissue integrity and RNA quality of laser microdissected phloem of potato. Planta 226, 797–803.

    Article  PubMed  CAS  Google Scholar 

  35. Asano, T., Masumura, T., Kusano, H., et al. (2002) Construction of a specialized cDNA library from plant cells isolated by laser capture microdissection: Toward comprehensive analysis of the genes expressed in the rice phloem. Plant J 32, 401–408.

    Article  PubMed  CAS  Google Scholar 

  36. Suwabe, K., Suzuki, G., Takahashi, H., et al. (2008) Separated transcriptomes of male gametophyte and tapetum in rice: Validity of a laser microdissection (LM) microarray. Plant Cell Physiol 49, 1407–1416.

    Article  PubMed  CAS  Google Scholar 

  37. Ishimaru, T., Nakazono, M., Masumura, T., et al. (2007) A method for obtaining high integrity RNA from developing aleurone cells and starchy endosperm in rice (Oryza sativa L.) by laser microdissection. Plant Sci 173, 321–326.

    Article  CAS  Google Scholar 

  38. Liu, H., Wang, S., Yu, X., et al. (2005) ARL1, a LOB-domain protein required for adventitious root formation in rice. Plant J 43, 47–56.

    Article  PubMed  Google Scholar 

  39. Klink, V., Alkharouf, N., Macdonald, M., and Matthews, B. (2005) Laser capture microdissection (LCM) and expression analyses of Glycine max (Soybean) syncytium containing root regions formed by the plant pathogen Heterodera glycines (Soybean Cyst Nematode). Plant Mol Biol 59, 965–979.

    Article  PubMed  CAS  Google Scholar 

  40. Klink, V., Overall, C., Alkharouf, N., Macdonald, M., and Matthews, B. (2007) Laser capture microdissection (LCM) and comparative microarray expression analysis of syncytial cells isolated from incompatible and compatible soybean (Glycine max) roots infected by the soybean cyst nematode (Heterodera glycines). Planta 226, 1389–1409.

    Article  PubMed  CAS  Google Scholar 

  41. Ithal, N., Recknor, J., Nettleton, D., Maier, T., Baum, T., and Mitchum, M. (2007) Developmental transcript profiling of cyst nematode feeding cells in soybean roots. Mol Plant-Microbe Interact 20, 510–525.

    Article  PubMed  CAS  Google Scholar 

  42. Sanders, P., Bui, A., Le, B., and Goldberg, R. (2005) Differentiation and degeneration of cells that play a major role in tobacco anther dehiscence. Sex Plant Reprod 17, 219–241.

    Google Scholar 

  43. Balestrini, R., Gómez-Ariza, J., Lanfranco, L., and Bonfante, P. (2007) Laser microdissection reveals that transcripts for five plant and one fungal phosphate transporter genes are contemporaneously present in arbusculated cells. Mol Plant-Microbe Interact 20, 1055–1062.

    Article  PubMed  CAS  Google Scholar 

  44. Ramsay, K., Wang, Z., and Jones, M. (2004) Using laser capture microdissection to study gene expression in early stages of giant cells induced by root-knot nematodes. Mol Plant Pathol 5, 587–592.

    Article  PubMed  CAS  Google Scholar 

  45. Kerk, N. M., Ceserani, T., Tausta, S. L., Sussex, I. M., and Nelson, T. M. (2003) Laser capture microdissection of cells from plant tissues. Plant Physiol 132, 27–35.

    Article  PubMed  CAS  Google Scholar 

  46. Tang, W., Coughlan, S., Crane, E., Beatty, M., and Duvick, J. (2006) The application of laser microdissection to in planta gene expression profiling of the maize anthracnose stalk rot fungus Colletotrichum graminicola. Mol Plant-Microbe Interact 19, 1240–1250.

    Article  PubMed  CAS  Google Scholar 

  47. Woll, K., Borsuk, L. A., Stransky, H., Nettleton, D., et al. (2005) Isolation, characterization, and pericycle-specific transcriptome analyses of the novel maize lateral and seminal root initiation mutant rum1. Plant Physiol 139, 1255–1267.

    Article  PubMed  CAS  Google Scholar 

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Acknowledgements

The authors would like to thank Bronwyn Carlisle for drawing Fig. 22.1. Protocols were developed in the Macknight laboratory with support from the Marsden Fund.

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Authors and Affiliations

  1. Department of Biochemistry, University of Otago, Dunedin, Otago, New Zealand

    Robert C. Day

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  1. Robert C. Day
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Editors and Affiliations

  1. (ETH), Department of Biology, Swiss Federal Institute of Technology, Universitätstrasse 2, Zurich, CH-8092, Switzerland

    Lars Hennig

  2. (ETH), Department of Biology, Swiss Federal Institute of Technology, Universitätstrasse 2, Zurich, CH-8092, Switzerland

    Claudia Köhler

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Day, R.C. (2010). Laser Microdissection of Paraffin-Embedded Plant Tissues for Transcript Profiling. In: Hennig, L., Köhler, C. (eds) Plant Developmental Biology. Methods in Molecular Biology, vol 655. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-60761-765-5_22

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  • DOI: https://doi.org/10.1007/978-1-60761-765-5_22

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-60761-764-8

  • Online ISBN: 978-1-60761-765-5

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