Re: [DIYbio] 4 months at -20

Thanks Koeng! Reading the first paragraph I was thinking "Survival's
fine, but what about plasmid?"..only to read further and see that you
have that covered, too! :)

Nice work, thank you for following up with the rest of us.

On 27/10/14 20:35, Koeng wrote:
> Hey!
>
> You might remember that a few months ago I posted on hypothetically
> storing E coli long-term at negative 20. After 4 months, I have verified
> that every condition I tried, 10%, 20% and 40% glycerol, all gave valid
> new colonies with the plasmid I had originally in them. It hasn't been a
> year yet, but this shows some promise for a technique DIYbiologists can
> use instead of storing DNA in plasmid form (I personally don't like
> doing that because its degraded for me before, or I don't have enough
> and the old never gets used, or I need a lot more and don't want to
> retransform)
>
> Experimental conditions (rough layout. Plasmid 7 is an RFP plasmid I use)
> 1 culture of plasmid 7, with 10% glycerol
> 1 culture of plasmid 7, with 20% glycerol
> 1 culture of plasmid 7, with 40% glycerol
>
> These 3 were placed in a frequently used -20 freezer box, and left for 4
> months.
>
> After 4 months, a small amount of liquid (~30µl) of each was put into
> new liquid culture, along with 1 from the -80 as a control (I should
> have done plates, sorry)
>
> After 16 hours of growth at 37c with shaking these were analyzed for RFP
> expression.
>
> RFP expression was confirmed. The cells, or at least a significant
> amount of the cells, had the plasmid and were growing.
>
>
> Richard Kramer has similar experience with this from the last thread
> - https://groups.google.com/forum/#!searchin/diybio/20$20freezer/diybio/lJjLfMX4OK4/G3VpcKEFTtwJ
> I also personally found that the cells tend to settle and that the
> liquid also stays viscus. I'll be continuing this experiment, but just
> wanted to give anyone that wanted to know a heads up that this is a
> viable way to store cells for at least 4 months, and reportedly 3 years.
> If anyone wants to try storing cells this way, or is storing them this
> way, give me a heads up so I can gather more data and replicates to give
> a definitive answer. Thanks!
>
> -Koeng
>
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