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Optimal Preparation of Tissue Sections for Light-Microscopic Analysis of Phloem Anatomy

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Phloem

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2014))

Abstract

In order to successfully analyze and describe any plant tissue, the first challenge is preparation of good anatomical slides. The challenge is even greater when the target tissue has heterogeneous characteristics, such as the phloem where soft and stiff tissues occur side by side. The goal of this chapter is to present a detailed protocol containing various techniques for optimal preparation of phloem tissue samples for light microscopic analysis. The process typically involves the steps of fixation, softening, embedding, sectioning, staining, and mounting. The protocol can be applied to make samples of phloem and surrounding tissues of stems and roots, from woody to herbaceous plants.

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References

  1. Grew N (1682) The anatomy of plants with an idea of a philosophical history of plants. Rawlins, London

    Google Scholar 

  2. Malpighi M (1686) Opera Omnia. Littleburry, London

    Google Scholar 

  3. Baas P (1982) Antoni Van Leeuwenhoek and his observation on the structure of the woody cell wall. IAWA 3:3–6

    Article  Google Scholar 

  4. Angyalossy V, Pace MR, Evert RF, Marcati CR, Oskolski AA, Terrazas T et al (2016) IAWA list of microscopic bark features. IAWA J 37:517–615

    Article  Google Scholar 

  5. Evert RF (2006) Esau’s Plant Anatomy: meristems, cells, and tissues of the plant body—their structure, function, and development. Wiley, Hoboken, NJ

    Book  Google Scholar 

  6. Sass J (1958) Botanical microtechnique. The Iowa State University Press, Ames

    Google Scholar 

  7. Johansen DA (1940) Plant microtechnique. McGraw-Hill Book Company, Inc, New York and London

    Google Scholar 

  8. Berlyn GP, Miksche JP (1976) Botanical microtechnique and cytochemistry. The Iowa State University Press, Ames

    Book  Google Scholar 

  9. Karnovsky MJ (1965) A formaldehyde-glutaraldehyde fixative of high osmolality for use in electron microscopy. J Cell Biol 27:137A–138A

    Google Scholar 

  10. Kukachka BF (1977) Sectioning refractory woods for anatomical studies. USDA Forest Service Research Note FPL-0236, Madison

    Google Scholar 

  11. Carlquist S (1982) The use of ethylenediamine in softening hard plant structures for paraffin sectioning. Stain Technol 57:311–317

    Article  CAS  Google Scholar 

  12. Rupp P (1964) Polyglykol als Einbettungsmedium zum Schneiden botanischer Präparate. Mikrokosmos 53:123–128

    Google Scholar 

  13. Barbosa ACF, Pace MR, Witovsk L, Angyalossy V (2010) A new method to obtain good anatomical slides of heterogeneous plant parts. IAWA J 31:373–383

    Article  Google Scholar 

  14. Bennet HS, Wyrick AD, Lee SW, McNeil JH (1976) Science and art in preparing tissues embedded in plastic for light microscopy, with special reference to glycol methacrylate, glass knives and simple stains. Stain Technol 51:71–97

    Article  Google Scholar 

  15. Hamann T, Smets E, Lens F (2011) A comparison of paraffin and resin-based techniques used in bark anatomy. Taxon 60:841–851

    Article  Google Scholar 

  16. Yeung EC, Chan CKW (2015) Glycol methacrylate: the art of embedding and serial sectioning. Botany 93:1–8

    Article  CAS  Google Scholar 

  17. Cheadle VI, Gifford EM, Esau K (1953) A staining combination for phloem and contiguous tissues. Stain Technol 28:49–53

    Article  CAS  Google Scholar 

  18. Bukatsch F (1972) Bemerkungen zur Doppelfärbung Astrablau-Safranin. Microkosmos 61:255

    Google Scholar 

  19. Kraus JE, Arduin M (1997) Manual básico de métodos em morfologia vegetal. Editora Universidade Rural, Seropédica

    Google Scholar 

  20. Chamberlain CJ (1935) Methods in plant anatomy. The University of Chicago Press, Chicago

    Google Scholar 

  21. O’Brien TP, Feder N, McCully ME (1964) Polychromatic staining of plant cell walls by toluidine blue. Protoplasma 59:368–373

    Article  Google Scholar 

  22. Ravikumar S, Surekha R, Thavarajah R (2014) Mounting media: an overview. J Dr NTR Univ Health Sci 3(Supplement 1):S1–S8

    Article  Google Scholar 

  23. Neuhaus B, Schmid T, Riedel J (2017) Collection management and study of microscope slides: storage, profiling, deterioration, restoration procedures, and general recommendations. Zootaxa 4322:1–173

    Article  Google Scholar 

  24. Loveland RP, Centifanto YM (1986) Mounting media for microscopy. Microscope 34:181–241

    CAS  Google Scholar 

  25. Pace MR, Lohmann LG, Angyalossy V (2011) Evolution of disparity between the regular and variant phloem in Bignonieae (Bignoniaceae). Am J Bot 98:602–618

    Article  Google Scholar 

  26. Pace MR, Alcantara S, Lohmann LG, Angyalossy V (2015) Secondary phloem diversity and evolution in Bignonieae (Bignoniaceae). Ann Bot 116:333–358

    Article  CAS  Google Scholar 

  27. Barbosa ACF, Costa GRO, Angyalossy V, Dos Santos T, Pace MR (2018) A simple and inexpensive method to sharpen permanent steel knives for microtomy. IAWA J 39:497–503

    Google Scholar 

  28. Cabanillas PA, Pace MR, Angyalossy V (2017) Structure and ontogeny of the fissured stems of Callaeum (Malpighiaceae). IAWA J 38: 49–66.

    Article  Google Scholar 

  29. Pace MR, Acevedo-RodrÚguez P, Amorim AM, Angyalossy V (2018) Ontogeny, structure and occurrence of interxylary cambia in Malpighiaceae. Flora 241: 46–60

    Article  Google Scholar 

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Acknowledgments

I would like to thank Antonio C. F. Barbosa from the Technological Research Institute of São Paulo for instructing me in microtomy, Gisele Costa from the University of São Paulo, Stan Yankowski for important inputs and for proofreading the manuscript, Mark Strong for proofreading the manuscript, Teresa Terrazas for Fig. 1b, the PeterBuck postdoctoral fellowship for funding at the Smithsonian Institution and the Programa de Apoyo a Proyectos de Investigación e Innovación Tecnológica (PAPIIT IA200319—Mexican government).

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Correspondence to Marcelo Rodrigo Pace .

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Pace, M.R. (2019). Optimal Preparation of Tissue Sections for Light-Microscopic Analysis of Phloem Anatomy. In: Liesche, J. (eds) Phloem. Methods in Molecular Biology, vol 2014. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9562-2_1

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  • DOI: https://doi.org/10.1007/978-1-4939-9562-2_1

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-9561-5

  • Online ISBN: 978-1-4939-9562-2

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