User:Justin M. Scott/Notebook/Col V Population Dynamics/2012/10/09

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Solid Media Experiment

10/9/2012 Methods:

  • Placed singles from an LB+CMP plate for BC208pACYC184 (Streaked 10/5/12) in 10mL of LB with 10µL of CMP (1000x), 10µL of AMP (1000x), and 10µL of STR (1000x)
  • Placed singles from an LB+NAL plate for Exp. Resist #2 (Streaked 10/5/12) in 10mL of LB with 10µL of NAL (1000x) and 10µL of AMP (1000x)
  • Placed singles from an LB+NAL+KN plate for BC212 (Streaked 10/5/12) in 10mL of LB with 10µL of NAL (1000x) and 20µL of KN (500x)
  • Incubated on roller drum at 37˚C and speed 5

10/10/2012 Method:

  • Diluted each O.N. to 107
  • Pipetted 30µL drops of diluted BC208pACYC184 onto LB plates in an circular patter, 8 drops to a circle
  • Allowed droplet to soak into the media
  • Pipetted 30µL drops of diluted Exp. Resist #5 onto LB plates in an circular patter, 8 drops to a circle
  • Allowed droplet to soak into the media
  • Pipetted 30µL drops of diluted BC212 onto LB plates in an circular patter, 8 drops to a circle
  • Allowed droplet to soak into the media
  • Conducted in triplicate
  • Incubated at 37˚C
  • After 24 hours of incubation, replica plated on LB+STR+AMP+CMP, LB+NAL+AMP, LB+NAL+KN plates and a new LB plate
  • Incubated at 37˚C for 24 hours
  • Replica plated the LB plate onto LB+STR+AMP+CMP, LB+NAL+AMP, LB+NAL+KN plates and a new LB plate
  • Added 1000µL of liquid LB to the LB+STR+AMP+CMP, LB+NAL+AMP, LB+NAL+KN plates
  • Scraped plate with a glass spreader
  • Poured off liquid LB into a sterile beaker on ice
  • Repeated the scrapping twice more (3000µL of liquid LB total for each plate)
  • Diluted the liquid in the sterile beaker to 108 (so a total dilution of 109)
  • Used droplet method for viable cell count onto new LB+STR+AMP+CMP, LB+NAL+AMP, LB+NAL+KN plates
  • Platted droplets in triplicate
  • Incubated all plates at 37˚C overnight
  • Replica plated the LB plate onto LB+STR+AMP+CMP, LB+NAL+AMP, and LB+NAL+KN plates
  • Placed the viable cell count plates in 4˚C refrigerator
  • Scrapped the LB+STR+AMP+CMP, LB+NAL+AMP, and LB+NAL+KN plates
  • Incubated all plates at 37˚C for 24 hours
  • Placed the viable cell count plates in 4˚C refrigerator
  • Scrapped the LB+STR+AMP+CMP, LB+NAL+AMP, and LB+NAL+KN plates
  • Incubated all plates at 37˚C for 24 hours
  • Placed the viable cell count plates in 4˚C refrigerator
  • Counted colonies on viable cell count plates

Results:

  • Solid Competition 10.10.12

Notes:

  • Replica platting 1 order: cmp,amp,kn
  • Master 3 fell out during replica platting
  • Replica platting 2 order: amp,kn,cmp
  • Replica 2 fell out during replica platting
  • Replica platting 3 order: kn,cmp,amp



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