You can use Agar-Tubes
fill one tube to 2/3 with steril LB add 0,6 % Agar and add max. 10 mg Cystein
pick one of your colony
and put it a few times in the agar
growth overnight at 37 °C
storage in darkness at 4 °C
renew it twice a year
Inoculate 4.5 ml of nutrient broth with bacteria. After incubation at 37 ° C overnight to obtain an optimum amount of bacteria, add 0.5 ml of DMSO and mix vigorously. Then place 1 ml aliquots den cryotubes The final concentration of the cryopreservative, glycerol or DMSO, should be at least 10%, v/v.
Una manera clásica de almacenar cualquier tipo de bacteria es suspenderla en leche desnatada adicionada de 5-10% de Glicerina (para que no se formen cristales durante la descongelación), hacer alícuotas y congelar. Para recuperarla, descongelar a 4ºC, resuspender por homogeinización y resembrar en los medios adecuados.
why dont to hand it over to culture collection centres like DSMZ for archiving at a low cost. for 1yr plus it would be good to freeze dry the sample in ampoules.
The best quality long-term maintenance of microbial cultures is certainly cryostorage in liquid nitrogen (culture medium containing 10% DMSO or 10% glycerol), if you have this possibility. Storage of yeast cultures at -20 C may be detrimental to them, better is routine passaging.
This is Protocol for Freezing Bacteria Using Glycerol.
You can use it.
Protocol for Freezing Bacteria Using Glycerol
Bacteria can be frozen using a solution of 15% glycerol. The process is simple and requires screw cap microfuge tubes and sterile glycerol. The glycerol is diluted to 30% so that it is easy to pipette. Equal amounts of 30% glycerol and culture broth are mixed, dispensed into tubes and then frozen.
Protocol
• Prepare a solution of 30% glycerol (v/v) by mixing 30 ml of glycerol with 70 ml of water. Transfer the solution to a screw cap glass bottle and sterilize by autoclaving at 121°C for 15 min. Loosen the cap during autoclaving.
• Aliquot 500 µl of sterile 30% glycerol into sterile 2 ml sterile microfuge tubes containing 4 mm glass beads*.
• Add 500 μl of bacterial culture to the tube and mix with the glycerol using a vortex mixer.
• Label the tube with the organism name, strain, date, etc.
• Place the tube in the freezer and record its location.
• To activate bacteria, only a couple of beads need to be removed from the tube. The tube doesn't need to be thawed. Open the tube and loosen a couple of beads using a sterile pipette tip. Pour the loose beads onto an agar plate and roll them around. Colonies that develop should be re-streaked. Replace the tube into the freezer immediately. If the culture thaws, do not re-freeze it as cells are typically very sensitive to freezing and thawing. Discard the thawed culture appropriately.
* We prefer tubes that are screw cap and have caps with O-rings. One option is to fill the tube 1/3 full with 4 mm glass beads (product number BAWG 4000-200-18) before sterilizing. When retrieving bacteria, remove or chip out one bead onto an agar plate and roll around to disperse the bacteria. This avoids thawing the entire stock.
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