Submitted by Lauren French (download pdf


 

Preparing a Mini Culture

Grow up a miniculture.

  1. Pipette 5.0 ml of LB into a 15ml polypropylene tube.
  2. Pipette 5.0 μl of antibiotic (50mg/ml for ampicillin) into tube.
  3. Using a sterile loop, add colony/colonies into the LB/antibiotic solution.
  4. Shake for 8 hours (or overnight) at 37°.


Making Glycerol Stocks

Make 4 glycerol stocks of the culture.

  1. Take 300μl of miniculture and add 700μl of LB + glycerol (1:1).
  2. Freeze all stocks at -20°; transfer two glycerol stocks to -80° after sitting at -20° overnight.


Preparing a Maxi Culture

  1. In a large flask, add 100 ml of LB and 100μl of antibiotic.
  2. Take between 200-500 μl of the miniculture and add to the flask.  If the miniculture did not grow well, use more volume of miniculture up to ~500μl.
  3. Shake for 16 hours or overnight.  Chill Sorvall GSA rotor in cold room overnight.

 

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