Journal article Open Access

Long Term Storage of Bacterial Strains

sprotocols


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  <identifier identifierType="DOI">10.5281/zenodo.13545</identifier>
  <creators>
    <creator>
      <creatorName>sprotocols</creatorName>
      <affiliation>ScientificProtocols.org</affiliation>
    </creator>
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  <titles>
    <title>Long Term Storage of Bacterial Strains</title>
  </titles>
  <publisher>Zenodo</publisher>
  <publicationYear>2014</publicationYear>
  <dates>
    <date dateType="Issued">2014-12-30</date>
  </dates>
  <resourceType resourceTypeGeneral="Text">Journal article</resourceType>
  <alternateIdentifiers>
    <alternateIdentifier alternateIdentifierType="url">https://zenodo.org/record/13545</alternateIdentifier>
  </alternateIdentifiers>
  <rightsList>
    <rights rightsURI="https://creativecommons.org/publicdomain/zero/1.0/legalcode">Creative Commons Zero v1.0 Universal</rights>
    <rights rightsURI="info:eu-repo/semantics/openAccess">Open Access</rights>
  </rightsList>
  <descriptions>
    <description descriptionType="Abstract">**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 &amp;quot; working&amp;quot; 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.</description>
  </descriptions>
</resource>
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