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Histostaining for Tissue Expression Pattern of Promoter-driven GUS Activity in Arabidopsis
Published: Jul 5, 2011 DOI: 10.21769/BioProtoc.93 Views: 31411
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Abstract
Promoter-driven GUS (beta-glucuronidase) activity is the most commonly used technique for tissue-specific expression patterns in Arabidopsis. In this procedure, GUS enzyme converts 5-bromo-4-chloro-3-indolyl glucuronide (X-Gluc) to a blue product. The staining is very sensitive. Processed samples can be examined under dissecting microscope or Differential Interference Contrast (Nomaski) microscope for bright blue color over cleared transparent background. Note this assay does not provide accurate information to subcellular levels.
Keywords: Gene expression
GUS activity
Histostaining
Arabidopsis
Promoter activity
Materials and Reagents
- Transgenic plants that contain genomic integration of a promoter: GUS expression cassette
- Potassium Ferrocyanide
- Potassium Ferricyanide
- Triton X-100
- 50 mM NaHPO4 buffer (pH 7.2)
- Dimethylformamide (DMF)
- Acetone
- NaHPO4 buffer
- 5-bromo-4-chloro-3-indolyl beta-D-glucuronide cyclohexamine salt (X-Gluc)
- 200 proof ethanol (once opened, 200 proof becomes essentially 190 proof)
- Staining buffer (see Recipes)
- Stock solutions (see Recipes)
Equipment
- Eppendorf tubes
- Vacuum
- Dissecting or light microscope
- Differential Interference Contrast (Nomaski) microscope
Procedure
Category
Cell Biology > Tissue analysis > Tissue staining
Plant Science > Plant cell biology > Tissue analysis
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