survival of frozen stocks

Doug Rhoads DRHOADS at MERCURY.UARK.EDU
Wed May 11 12:16:06 EST 1994


>To:             yeast at net.bio.net
>From:           stearns at leland.Stanford.EDU (tim stearns)
>Subject:        Re: survival of frozen stocks
>Date sent:      11 May 1994 15:19:10 GMT

>In article <BRATTY.94May10181611 at med04.bch.umontreal.ca>,
>Bratty John <bratty at BCH.UMontreal.CA> wrote:
>>
>>I made the cells 20% glycerol in a volume of 50 ul and flash-froze
>>them in a dry ice - ethanol bath.  After about 15 min, I thawed them
>>on ice, made serial dilutions and plated on YPD.  The control,
>>obviously, was an aliquot made 20% glycerol, but just left on ice
>>while the experimental tube was in the dry ice bath.
>>
>>To my surprise and dismay, when I counted the colonies, I found that
>>the mortality exceeded 99.99%, i.e. a four order of magnitude drop in
>>the number of viable cells after one such cycle of freezing and
>>thawing.
>
>I don't know if it makes a difference, but what I have always done is
>to make the cells 25% glycerol and freeze by putting in the -70
>freezer directly.  Although I haven't quantitated viability there is
>no way that we are getting a four order of magnitude drop from one cycle
>of freezing.  I don't think the flash-freezing is a good idea.  
>
>Tim Stearns
>stearns at leland.stanford.edu
>
We have been using 40% glycerol for our freezing of yeasts and bacteria.  
WE have not evaluated survival frequencies but have usually had good 
growth even after several freeze/thaw cycles.  Freezing is by placing into 
-85 without dry-ice/ethanol.

Doug Rhoads                  || Dept. of Biological Sciences
drhoads at mercury.uark.edu     || 601 Science Engineering
drhoads at uafsysb.uark.edu     || University of Arkansas
501-575-3251                 || Fayetteville, AR 72701



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