Bacterial glycerol stock is a type of suspension used in laboratory settings for long-term storage of plasmids. The bacteria on an LB agar plate can live for at least 2 weeks when stored at 4°C. You should plan to create a glycerol stock for long-term storage. When liquid bacteria cultures are mixed with a 50% glycerol solution, it will stabilize and protect the cell membrane from deterioration and temperature-related damage. A glycerol stock of bacteria can be stored stably at -80°C for extended periods of time. Note：Incubation conditions including growth time and growth temperature depend on different strains, which can be found on your plasmid's information page.
2. Add 500 μL overnight culture to 500 μL 50% glycerol in a 2 mL screw top tube or cryovial.
Stir or shake the container thoroughly until the liquids are evenly mixed.
Note：The optimal concentration of glycerol is undetermined, some labs prefer to use a glycerol solution as low as 15-40% in order to avoid compromising the bacterial culture.
Note：If possible, use a tube with a screw top rather than one with a snap top as they come open accidentally during storage.
3. Mark key information on the tube where it will be plainly visible.
Note: Be sure to record useful contents including the name of each part, the carrier name, the strain name, operation date, name of personnel, etc.
4. Place the labelled tubes at -80°c. Most plasmids are stable for years when stored as bacterial glycerol stocks.
Note: Repeated freeze and thaw cycles will reduce its lifespan considerably.
5. When it comes time to revive your stocks, use a sterile inoculation loop or pipette tip to scrape some bacteria off the top and streak the bacteria onto an LB agar plate for analysis.
Note: Operate as possible as quickly to avoid the glycerol stock unthaw.
6. Inoculate the single colony of bacteria or liquid culture as normal.

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