IGEM:PennState/2006/Motility-assays

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Contents

Swarm Plate Assays

Swarm Plate Assays

Time: 2 hr first day; 9 hr second day

Reference: Garza, A.G., et al. J. Mol. Biol. (1996) 258, 270-285

Media/Reagents/Materials

  • LB
  • Typtone broth plates
  • Cells (from plate)

Day 1: Make Tryptone Swarm Plates

1. For each plate to be made, add the following to a 125mL flask:

  • 0.25g Bacto tryptone
  • 0.20g NaCl
  • 0.15g sodium citrate
  • 0.05g Eiken agar (for a 0.20% agar swarm plate)

2. Add 25mL purified water to the flask and swirl to dissolve

3. Autoclave 25 minutes with slow exhaust

4. Upon cooling, add appropriate antibiotics and inducers

  • IPTG saturation concentration => 1mM
  • HSL saturation concentration => 100µM

5. Allow plates to dry overnight

Day 1: Inoculate O'N Culture

1. Inoculate 3mL overnight cultures of all samples to be tested

2. Shake at 200 RPM in 30°C incubator

Day 2: Assay

1. Dilute overnight culture 1:100 in LB (30uL of culture in 3mL LB)

2. Grow in a 30°C incubator at 200 RPM to an A590nm O.D. between 0.3-0.4

3. Add about 3uL of each culture to plates

4. Grow in a 30°C incubator 8 hours

5. Take picture of plate with digital camera

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