Wiesner Staining for Visualizing Lignin: A Technique to Stain Lignin Deposits Present in Stems of Arabidopsis Thaliana
Published: April 30, 2023
Abstract
Source: Pradhan Mitra, P. et al., Histochemical Staining of Arabidopsis thaliana Secondary Cell Wall Elements. J. Vis. Exp. (2014).
In this video, we describe the Wiesner staining technique for staining the lignin present in specialized plant cells. This staining technique helps to visualize the lignified cell walls in Arabidopsis thaliana stems and helps to quantify the lignification in the cell walls of different types of cells.
Protocol
1. Stem Embedding Make a 7% agarose solution in water (7 g of electrophoresis-grade agarose in 100 ml distilled water). Dissolve the agarose by autoclaving for 20 min or by microwaving for 20 min at lowest intensity (e.g., 10% intensity of a 1,250-watt microwave). Prepare a homemade mold for embedding the stems using plastic vials. Using a razor blade, cut off the conical bottom of a 2 ml screw-cap microcentrifuge tube (Part A). With a syringe needle, puncture a hole in the…
Representative Results
Figure 1. Homemade mold for embedding the stems. On the left side; top of the 2 ml screw-cap microcentrifuge tube (Part A) and the bottom from the 0.6 ml microcentrifuge tube (Part B). On the right side; the Parafilm holding the two parts; A and B to form the final mold. Figure 2. Phloroglucinol-HCl sta…
Declarações
The authors have nothing to disclose.
Materials
| Agarose | EMD | MERC2125 | CAS Number: 9012-36-6 |
| Phloroglucinol | Sigma | P 3502 | 1,3,5-trihydroxybenzene [CAS Number: 108-73-6] |
| Hydrochloric acid | EMD | HX0603-75 | CAS Number: 7647-01-0 |
| Ethanol 190 proof | KOPTEC | V1401 | CAS Number: 64-17-5 |
| Vibratome | Leica | Leica Vibrating blade microtome VT1000S | http://www.leicabiosystems.com/ products/sectioning/vibratingblade-microtomes/details/product/ leica-vt1000-s/ |
| Razor | American Safety razor company | Item # 60-0139-0000 | Stainless Steel Double Edge Blade (Personna Super) |
| Screw Cap Microcentrifuge Tubes (2 ml) | VWR | 16466-044 | |
| Microcentrifuge Tubes (0.6 ml) | Axygen Scientific | MCT-060-C | |
| Tissue adhesive | Ted Pella Inc | 10033 | Store at 4 °C or 20 °C for 3 months or longer storage |
| Camera with CCD chip with no mechanical shutter | Hamamatsu | C4742-95 | |
| Camera software | Molecular Devices | MetaMorph version 7.7.0.0 | |
| Imagining anaylsis | Adobe | Photoshop CS4 | |
| Micro Cover Glasses, Square, No. 1 | VWR | 48366-067 | 22 x 22 mm (7/8 x 7/8")-Cover glasses are corrosion-resistant |
| Frosted Micro Slides, 1 mm | VWR | 48312-003 | 75 x 25 mm – 1 mm |
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Wiesner Staining for Visualizing Lignin: A Technique to Stain Lignin Deposits Present in Stems of Arabidopsis Thaliana. J. Vis. Exp. (Pending Publication), e21057, doi: (2023).