Reagent Recipes

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

1. Our catalase is 13,000U/mg. We want a final concentration of 30,000U / ml. Therefore, to make 10ml, we need to use 23.07mg, or 0.0231g.
2. Use the precise balance of Karl's lab to measure out the catalase. Use a folded piece of weighing paper instead of a weighboat.
3. Put catalase into a 15ml sterile plastic tube.
4. Pipet in PBS, measuring out the volume using a serological pipet. Normally, we would use 10ml.
5. Gently shake until dissolved. DO NOT VORTEX.
6. Sterilely filter solution using a syringe filter and place into a fresh, sterile plastic tube.
7. Label top with contents and date; limit exposure to light by creating a aluminum foil sleeve.
8. Keep in refrigerator