Glycerol Stocks for Long-Term Storage of Bacteria

Notes

• A common technique to preserve E. Coli containing specific plasmids (and other bacterial) longer term.
• While it is possible to make a long term stock from cells in stationary phase, ideally your culture should be in the logarithmic growth phase.

Source

Standard lab technique – handed down from Postdoc to student.

Materials

• 1:1 glycerol/water solution (preferably be autoclaved)
• Cryogenic vials or (1.5 – 2µL) Eppendorf tubes – Screw caps preferred if snap-freezing in liquid N₂ to avoid the risk of the lids coming off.
• LN₂ – if at hand, cells are still recoverable even without, but slow-freeze can rupture the cells and reduces viability.

Method

1. Add 0.7 – 1 ml of 50% glycerol in H₂O to a tube.
2. Add 0.7 – 1 ml sample from the culture of bacteria to be stored.
3. Gently vortex the tube to ensure the culture and glycerol is well-mixed. Alternatively, pipet to mix.
4. Use a tough spot sticker to put the name of the strain or some useful identifier on the top of the vial.
5. On the side of the vial list all relevant information – part, vector, strain, date, researcher, etc.
6. Snap-freeze in LN₂ – if possible, does work without.
7. Store in a freezer box in a -80°C freezer. Remember to record where the vial is stored for fast retrieval later.