Reagent Recipes: Difference between revisions
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PreDec2022>EricMiller No edit summary |
PreDec2022>EricMiller No edit summary |
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=Reagent Information= | ==Reagent Information== | ||
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=Glycerol for freezing aliquots= | ==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%. | 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%. | ||
=10x PBS= | ==10x PBS== | ||
Used to create 1x PBS for dilution or washing cells of any species. | Used to create 1x PBS for dilution or washing cells of any species. | ||
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=Tris Buffer= | ==Tris Buffer== | ||
For Tris buffer at pH 8.0 (at 25 degrees) | For Tris buffer at pH 8.0 (at 25 degrees) | ||
*4.44 g/L Tris HCl | *4.44 g/L Tris HCl | ||
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'''Do not adjust pH or check the pH with the MEE lab pH probe. This will destroy the probe!''' | '''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. | |||
# Sterilely filter solution using a syringe filter and place into a fresh, sterile plastic tube. | |||
# Label top with contents and date; cover with foil to prevent exposure to light. | |||
# Keep in refrigerator | |||
Revision as of 16:01, 11 March 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 |
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%.
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.
- Sterilely filter solution using a syringe filter and place into a fresh, sterile plastic tube.
- Label top with contents and date; cover with foil to prevent exposure to light.
- Keep in refrigerator