Reagent Recipes: Difference between revisions
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PreDec2022>DavisonHammond No edit summary |
PreDec2022>EricMiller |
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Line 25: | Line 25: | ||
| 1000x | | 1000x | ||
| Light sensitive. Molecular weight: 238.31 g/mol | | Light sensitive. Molecular weight: 238.31 g/mol | ||
|- | |||
| Catalase | |||
| Cat | |||
| 30,000U / ml | |||
| H2O | |||
| 100x | |||
| Light sensitive. Do not vortex when dissolving. | |||
|- | |||
| NAD(+) / Factor V | |||
| NAD(+) | |||
| ???? | |||
| ???? | |||
| ???? | |||
| ???? | |||
|- | |||
| Hemin / Factor X | |||
| Hemin | |||
| ???? | |||
| ???? | |||
| ???? | |||
| ???? | |||
|- | |||
| Ethylhydrocupreine hydrochloride | |||
| Optochin | |||
| ??? | |||
| ??? | |||
| 1:100,000 final concentration | |||
| ???? | |||
|} | |} | ||
Revision as of 09:21, 17 October 2019
Reagent Information
Reagent | Abbreviation | Stock Solution
Concentration |
Solvent | Stock Concentration to
Working Concentration |
Notes |
---|---|---|---|---|---|
Bromo-chloro-indolyl-galactopyranoside | X-gal | 20mg/ml | Dimethylformamide (DMF) | 250x | Use DMF in a chemical hood |
Isopropyl β-D-1-thiogalactopyranoside | IPTG | 0.1M | H2O | 1000x | Light sensitive. Molecular weight: 238.31 g/mol |
Catalase | Cat | 30,000U / ml | H2O | 100x | Light sensitive. Do not vortex when dissolving. |
NAD(+) / Factor V | NAD(+) | ???? | ???? | ???? | ???? |
Hemin / Factor X | Hemin | ???? | ???? | ???? | ???? |
Ethylhydrocupreine hydrochloride | Optochin | ??? | ??? | 1:100,000 final concentration | ???? |
Glycerol for freezing aliquots
We use sterile 80% glycerol in water. We dilute this 1:3 (400ul 80% glycerol; 1200ul culture) for creating freezer stocks with a final glycerol concentration of 20%.
MgCl2
When working with MgCl2 be aware that when added to water a significant exothermal reaction occurs.
10x PBS
Used to create 1x PBS for dilution or washing cells of any species.
For 1 liter:
- NaCl 80g
- KCl 2g
- Na2HPO4 14.4g
- KH2PO4 2.4g
Stir salts with MilliQ water. This may take a significant time to dissolve!
Adjust pH with NaOH to 7.4. This will take a significant amount of NaOH; add in 200ul increments.
Tris Buffer
For Tris buffer at pH 8.0 (at 25 degrees)
- 4.44 g/L Tris HCl
- 2.65 g/L Tris Base
Do not adjust pH or check the pH with the MEE lab pH probe. This will destroy the probe!
Catalase
- Calculate the amount of catalase needed; final concentration should be 30,000U / ml. Each batch may require a different number of milligrams of catalase.
- Use the precise balance of Karl's lab to measure out the catalase. Use a folded piece of weighing paper instead of a weighboat.
- Put catalase into a 15ml or 50ml plastic tube.
- Pipet in PBS, measuring out the volume using a serological pipet.
- Gently shake until dissolved. DO NOT VORTEX.
- Sterilely filter solution using a syringe filter and place into a fresh, sterile plastic tube.
- Aliquot 1 ml in 1.5 ml microcenterfuge tubes.
- Label top with contents and date; limit exposure to light.
- Keep in refrigerator