Mar 12, 2026
Preparing a Glycerol Stock
This protocol is a draft, published without a DOI.
- Tamra Lahom1
- 1Carleton College
Protocol Citation: Tamra Lahom 2026. Preparing a Glycerol Stock. protocols.io https://protocols.io/view/preparing-a-glycerol-stock-jv2kcn8cx
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: March 12, 2026
Last Modified: March 12, 2026
Protocol Integer ID: 313132
Keywords: glycerol stock of bacteria, glycerol stock bacterial glycerol stock, glycerol stock, term storage of plasmid, frozen bacteria, bacteria on an lb agar plate, glycerol, addition of glycerol, bacteria
Abstract
Bacterial glycerol stocks are important for long-term storage of plasmids. Bacteria on an LB agar plate can be stored at 4°C for a few weeks. However, if you want to store bacteria for a longer time, you will need to establish glycerol stocks. The addition of glycerol stabilizes the frozen bacteria, preventing damage to the cell membranes and keeping the cells alive. A glycerol stock of bacteria can be stored stably at -80°C for many years.
Troubleshooting
Label the tube with the date, the glycerol stock number, the protein of interest, and the plasmids or antibiotics
Note
You need to label before flash freezing so the maker appears on the tube.
Combine 500 µL of overnight with 500 µL of 50% Glycerol
Note
This needs to be performed under sterile conditions so under a flame
Flash freeze in Liquid Nitrogen then store in the -80 C.