Journal article Open Access

Long Term Storage of Bacterial Strains

sprotocols


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  <dc:creator>sprotocols</dc:creator>
  <dc:date>2014-12-30</dc:date>
  <dc:description>**Time required:**

Cells are grown 6 hours to overnight; a total of less than 5 minutes bench time for each strain.

### Procedure:

**Day 1**

1. Inoculate a 15 ml culture tube containing 5 ml of LBM or LBM+antibiotic selective medium with a freshly grown isolated colony. Incubate at 37 degrees C until culture is in late log or stationary phase (usually 5 hours to overnight).


**Day 2**

1. For each strain to be stored at -80 degrees C for the archives prepare a sterile labeled cryovial. Pipet 225 ul sterile 80% glycerol into the cryovial. Add 1.0 ml of the bacterial culture (frozen stock will be 15% glycerol). Mix well (vortex) and place tube at -80 degrees C.
- For each strain to be stored at -20 degrees C as a liquid glycerol " working" stock pipet equal volumes 80% glycerol and bacterial culture into a labeled polypropylene tube. Mix the contents well (if not well mixed ice crystals will form decreasing the viability of the cells). Place the tube in a -20 degrees C freezer. If possible check the viability of the cells after 1 week.


To recover a strain from the -80 degrees C glycerol stock use a sterile toothpick to scrape some of the ice then streak out the cells on the appropriate medium e.g. LBM + ampicillin. Do not thaw the frozen stocks because each freeze-thaw cycle will result in a 50% loss in cell viability.

To use the -20 degrees C working stocks pipet 50 to 100 ul as inoculum for a 5 ml overnight culture.</dc:description>
  <dc:identifier>https://zenodo.org/record/13545</dc:identifier>
  <dc:identifier>10.5281/zenodo.13545</dc:identifier>
  <dc:identifier>oai:zenodo.org:13545</dc:identifier>
  <dc:rights>info:eu-repo/semantics/openAccess</dc:rights>
  <dc:rights>https://creativecommons.org/publicdomain/zero/1.0/legalcode</dc:rights>
  <dc:title>Long Term Storage of Bacterial Strains</dc:title>
  <dc:type>info:eu-repo/semantics/article</dc:type>
  <dc:type>publication-article</dc:type>
</oai_dc:dc>
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