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Indole Derivative Feeding Test and Detection of TRP and Indole derivatives by Thin Layer Chromatography

吲哚衍生物的喂养试验及薄层色谱法检测TRP和吲哚衍生物

MH Magdalena Hilbert
LV Lars M. Voll
JH Jörg Hofmann
AZ Alga Zuccaro

发布: 2013年06月20日第3卷第12期 DOI: 10.21769/BioProtoc.801 浏览次数: 11345

评审: Tie Liu

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Abstract

The mutualistic root endophyte Piriformospora indica colonizes a wide range of plants and the colonization of root cells by this fungus is very often associated with beneficial effects to its host, such as growth promotion and increased biotic and abiotic stress tolerance. These traits could be based on general mechanisms and signaling pathways common to many different plant species. One such mechanism could be the recruitment of phytohormone pathways by P. indica. It is known, that many mutualistic microorganisms are able to synthesize and secrete phytohormones during the interaction with their host plants. This protocol has been successfully utilized to analyze tryptophan (TRP)-dependent biosynthesis of indole-3-acetic acid (IAA) and its indole derivatives by P. indica (Hilbert et al., 2012).

Materials and Reagents

  1. Indole derivative:
    TRP (Sigma-Aldrich, catalog number: T0254-500g )
    IAD (indole-3-acetaldehyde) (Sigma-Aldrich, catalog number: I1000-100mg )
    IAA (Sigma-Aldrich, catalog number: I5148-2g )
  2. Standard microscope coverslips
  3. Standard microscope slides
  4. 0.002% (v/v) Tween water 20
  5. 0.9% NaCl
  6. FeCl3
  7. HClO4
  8. NaNO3
  9. MgSO4.7H2O
  10. KH2PO4
  11. Orthophosphoric acid
  12. Glucose
  13. Peptone
  14. Yeast extract
  15. Casamine acids
  16. Microelements
  17. Agar
  18. Tryptophan (Sigma-Aldrich, catalog number: T0254-500g)
  19. Ethyl acetate
  20. Aluminum foil
  21. p-dimethylaminobenzaldehyde (Sigma-Aldrich)
  22. Van Urk reagent (see Recipes)
  23. Salkowski test reagents (see Recipes)
  24. 10 mM orthophosphoric acid (see Recipes)
  25. Complete medium (see Recipes)
  26. 20x salt solution (see Recipes)
  27. Microelements (see Recipes)

Equipment

  1. Erlenmeyer flasks (100 ml)
  2. Neubauer improved counting chamber (Marienfeld-Superior, Lauda, Königshofen, Germany)
  3. Scalpel (sterile)
  4. Whatman paper
  5. Miracloth filter 15 cm x 15 cm (Merck KGaA, catalog number: 475855 )
  6. Drigalski spatula (sterile)
  7. Disposable cuvettes
  8. Preval sprayer
  9. Thin layer chromatography (TLC) chamber
  10. Petri dishes (12 mm in diameter)
  11. 15 ml sterile Falcon tubes
  12. 2 ml Eppendorf tubes
  13. Glass test tubes
  14. Incubator e.g. Certomat BS-1 (B. Braun Biotech International)
  15. Speedvac sc110 (Savant, Thermo Fisher Scientific)
  16. Biofuge Primo R (Heraeus) (Rotor, catalog number: 7590 )
  17. Heraeus Pico 17 centrifuge (Heraeus)
  18. Spectrophotometer e.g. Ultrospec 3,000 proUV/Visible (GE Healthcare)
  19. Clean bench Hera Safe (Heraeus)
  20. Hybridization oven (Hybaid Shake n’ Stack, Thermo Fisher Scientific)

Procedure

English
中文翻译

文章信息

版权信息

© 2013 The Authors; exclusive licensee Bio-protocol LLC.

如何引用

Hilbert, M., Voll, L. M., Hofmann, J. and Zuccaro, A. (2013). Indole Derivative Feeding Test and Detection of TRP and Indole derivatives by Thin Layer Chromatography. Bio-protocol 3(12): e801. DOI: 10.21769/BioProtoc.801.

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分类

植物科学 > 植物生理学 > 胞内共生

微生物学 > 微生物-宿主相互作用 > 真菌

生物化学 > 其它化合物 > 植物激素 > 乙烯

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