Mike Barnkob:Protocols/Bacterial/ON liquid culture

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Inoculating overnight liquid culture

Pick colonies and inoculate a liquid culture.

Reagents

Prepare liquid LB media:

• Following into a 500 ml flask:
  8.2 g LB Broth Ezmix (which includes Tryptone, 5 g/L Yeast Extract, 5 g/L NaCl)
  dH2O up to 400mL
• Cover flask with aluminum foil, label with autoclave tape and autoclave.
• Add ampicillin to a concentration of 100 µg/mL (0.4 ml of 100mg/mL)

Protocol

For a single colony:

1. Add 5 ml of LB + antibiotic to two 50ml Falcon tubes.
2. Using a sterile loop, select a single colony from LB plate and dip into Falcon Tube.
3. Loosely cover the culture with sterile aluminum foil or a cap that is not air tight.
4. Incubate overnight (12-18 hours) in shaking incubator at 37°C.
   Note: Also incubate a tube that only has LB + antibiotic, but no bacteria as a control. No bacteria should be visible in this after incubation.
5. Check for growth
6. Proceed to create a glycerol stock and isolate plasmid DNA.

References

• http://www.addgene.org/recipient-instructions/inoculate-bacterial-culture/