Grow an overnight culture in M9 Media

• Aliquot 5 mL M9 media into an overnight tube
• Add Antibiotics (5uL Amp 100)
• Inoculate desired strain
• Incubate and shake at 200rpm, 37 C

Grow the overnight culture in exponential phase

• Aliquot 5 mL M9 media into an overnight tube
• Add Antibiotics
• Add 50 uL of overnight culture (shake gently to mix the cells that may have pelleted) into fresh media
• Incubate and shake at 200 rpm, 37C
• After 3 hours, test OD with cuvettes to check for exponential growth (usually around 0.3)
• In 15 min intervals, check OD to make sure cells are in exponential phase

Prepare plate (should be done during the incubation time for growing exponential cells)

• Add 175 uL of desired media (if testing with pH media, add it now)
• Record which well contains what media
• Make sure to work in triplicates

Plating exponential cells

• Add 1 uL of exponential cell culture into desired well
• Place in plate reader and test for both absorbance 600 and fluorescence 485
• Test for 16 hours (time can be adjusted)