THIN LAYER CHROMATOGRAPHY |
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Last Update: December 2006 |
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References: Bonin CP, Potter I, Vanzin GF, Reiter WD (1997) The MUR1 gene of Arabidopsis thaliana encodes an isoform of GDP-D-mannose-4,6-dehydratase, catalyzing the first step in the de novo synthesis of GDP-L-fucose. PNAS 94: 2085-2090; Orellana A, Neckelmann G, Norambuena L (1997) Topography and function of Golgi uridine-5'-diphosphatase from pea stems Plant Physiology 114: 99-107 |
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| PREPARE SOLUTIONS | |
| 1. Acetic acid (glacial): | Commercial acetic acid (glacial) is 17.4N |
| 2. 3M LiCl (1 L): | Mix 127.17 g of LiCl with 1 L of dH2O |
| 3. 1 M Ammonium acetate (1 L): | Mix 77.08 g of ammonium acetate with 1 L of dH2O |
| PROCEDURE - Acetic acid: lithium chloride TLC | |
| 1. Pre-run with 0.3M LiCl all ~20 cms of the plate | |
| 2. Dry plate (>1 hr) | |
| 3. Load samples (as close to 1 mL as possible) and let them dry | |
4. Run for 2 cms with 1 N acetic acid |
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| 5. Dry (>1 hr) | |
| 6. Run the rest of the plate (~15 cms) with a mix of 9 volumes of 1N acetic acid with 1 volume of 3M LiCl | |
| PROCEDURE - Ethanol: ammonium acetate TLC | |
| 1. Mix 7 vols of 100% ethanol with 3 vols 1M ammonium acetate, pH 7 (can be pH as low as 3.8) | |
| NOTE: instead of 7 vols of 100% ethanol, 5 volumes can be tested | |
2. Run until front reaches top of plate |
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| PROCEDURE - 1-butanol:acetic acid:water TLC (12:3:5) | |
| 1. Mix in the above volume ratios 100% 1-butanol: 1N acetic acid: water | |
| 2. Run until the front reaches the top of the plate | |
| PROCEDURE - Acetate:pyridine:water TLC (8:2:1) | |
| 1. Mix in the above volumes | |
| 2. Run until the front reaches the top of the plate | |
| PROCEDURE - 1-butanol: pyridine: water (6:4:3) | |
| 1. Mix in the above volumes | |
| 2. Run until the front reaches the top of the plate | |