Prerequisite Protocols

• None

Materials

• MES Monohydrate (10mM final)
• Magnesium Chloride Hexahydrate (10mM final) (or other hydrations; adjust to match final desired molarity)
• Potassium Hydroxide 1M (for pH adjustment)
• 0.22um Filter Membrane, or syringe filter if you feel strong
• Alpha Lipoic Acid 1000x Stock (5uM final) (added ONLY at time of use)
• Acetosyringone (100uM final) (added ONLY at time of use)

Equipment

• Magnetic Stirrer
• pH Meter (freshly calibrated)
• Vacuum Filtration Apparatus (or large syringe)
• 4C Refrigerator

Step By Step

1. To a 500mL beaker, add 450mL of distilled water and begin stirring.
2. Weigh out 1.017g of Magnesium Chloride Hexahydrate (203.30g/mol) and add it to the stirring water.
3. Weigh out 1.067g of MES Monohydrate (213.3g/mol) and add it to the stirring water.
4. Calibrate pH meter and adjust pH of stirring buffer to 5.60 using 1M KOH via a 200uL pipette. Go slowly and do not overshoot. pH will rise quickly until reaching the buffing zone of 5.4ish. Allow a few seconds between additions of KOH for pH reading to stabilize.
5. Dispense the pH adjusted buffer into a clean 500mL volumetric flask and adjust volume to the 500mL neck mark.