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If I kept E. coli transformed with plasmid at -20˚C without making glycerol stock, will it survive? Actually I have kept it this way for 18 hours. What will happen with it? Will it grow in fresh media?

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My guess is that you may be able to rescue something at this stage, but this is definitely not recommended for long-term storage. If you can't recover any living cells you should still be able to recover some plasmid from the cells for retransformation. – Alan Boyd Sep 18 '12 at 16:45
e coli are tough bugs. they will last in the freezer for a while, but good experimentalists don't leave things like this to chance. They live fine in a regular refrigerator for a week or more - no need to freeze. replate and then transfer to glycerol asap. – shigeta Sep 18 '12 at 21:46

2 Answers

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Trehalose[1],[2] is the cryoprotectant used by bacteria to keep things going when all else freezes (or dries up), and they grow in it, too!

Trehalose accumulates dramatically in microorganisms during heat shock and osmotic stress and helps protect cells against thermal injury and oxygen radicals. Here we demonstrate an important role of this sugar in cold-adaptation of bacteria.

  1. Kandror O, DeLeon A, Goldberg AL. 2002. Trehalose synthesis is induced upon exposure of Escherichia coli to cold and is essential for viability at low temperatures. Proceedings of the National Academy of Sciences USA, 99(15): 9727-9732, doi:10.1073/pnas.142314099
  2. http://en.wikipedia.org/wiki/Trehalose
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Over time I suspect that ice crystals will form in the colonies from defrost cycles in the freezer and the moisture in the agar. This will cause rupture of the bacteria membranes and over time may destroy the bulk of the bacteria cells. The plates will also desiccate over time as well from the defrosting cycles of the freezer. You may be able to leave them in the freezer for a while but I can't foresee them surviving indefinitely.

If the strains are really important I recommend just sucking it up and spending the small amount of time necessary to make the glycerol stock. Glycerol is used in bacteria stocks as a cryoprotectant to prevent the crystallization of the stock.

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