Freezing Bacterial Stocks

Freezing Bacterial Stocks

- Good microbiologists always start with a single colony, so streak a plate (with appropriate antibiotic) using standard isolation technique (streak; flame; streak; flame; streak...look it up).

- For freezing, do one of the following: 

A) Streak a single colony on a plate (with antibiotic).  The next morning, Put ~ 1 ml of 50% LB 50% Glycerol (sterile, of course) into a sterile cryovial (like the ones we use to freeze ES cells).  Scrape 3 or 4 loopfuls of bacteria off the plate and disperse into the freeze medium.  Make sure to mix thoroughly by vortexing and/or pipetting.

B) Innoculate 3 ml of LB (with antibiotic) from a single colony on on your isolation plate.  Grow several hours-overnight.  Mix 0.5ml with 0.5ml of sterile glycerol. 

- Put the vial into one of the Lab’s Bacterial Strain Boxes in the -80.  Make sure you add it to the most current box.

- Go to our Filemaker database on the shared iMAC, open the Frozen Bacterial Stocks database (there is an alias icon on the desktop, when logged in for General Lab Use), and enter the information in the fields.  Importantly, put the correct box/slot numbers.  

- TO RETRIEVE FROZEN CLONES:  Get out the vial. Before it fully thaws, scrape the top with a sterilized loop, and streak a plate for isolation.  Put the vial back in -80. 

Questions?  Contact John at