Mike Barnkob:Protocols/Bacterial/LB agar plates

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Preparing LB plates with ampicillin

Recipe for preparing LB ampicillin plates.

Reagents

Weigh out the following into a 0.5L flask:

  • LB Broth Ezmix, 14.2mg
  • dH20, 400 ml

Note: Makes about 25 plates.

Protocol

  1. Mix LB broth mix into water by swirling
    Note: powder left on the sides of the flask will caramelize on the glass during autoclaving.
  2. Cover flask with aluminum foil and label with autoclave tape.
  3. Autoclave on the liquid setting for 20 minutes
  4. Allow the agar solution to cool to 55°C by placing in 55°C water bath.
  5. Add ampicillin to a concentration of 100 µg/mL (0.4 ml of 100mg/mL).
  6. Pour ~20mL of LB agar per 10cm polystyrene Petri dish
    Note: Before pouring, burn the flask head with a Bunsen burner.
    Note: Pour slowly from the flask into the center of the petri dish. When the agar has spread to cover about 2/3 of the dish stop pouring and the agar should spread to cover the entire plate.
  7. Place the lids on the plates and allow them to cool for 30-60 minutes. Let sit for several more hours or overnight.
  8. Label the bottom of plates with antibiotic and date and store in plastic bags or sealed with parafilm at 4°C.
    Note: Plates should be good for 1-2 months.

References

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