Re: [DIYbio] Freezing cells with plasmids

On Fri, Jun 27, 2014 at 7:33 PM, John Griessen <john@industromatic.com> wrote:
> On 06/27/2014 06:06 PM, Koeng wrote:
>>
>> Has anyone tried this?
>
>
> No, but I thought it was the rapidity of freezing that avoided
> cells being stabbed/deformed by ice crystals by way of smaller ice crystals
> in rapid freezing... If so, freezing in your -80, or liquid N2, then
> storing in -20 might get results.
>

I thought that was what the glycerol was for. Ah, glycerol is a
cryoprotectant [1]. It prevents the formation of ice crystals. My
procedure is ~350uL glycerol (eyeball it in a 2mL microcentrifuge
tube) then adding 500mL fresh bacteria culture. I quickly vortex until
fully mixed and stick in the -80C. I just cultured some recently that
I stored away last year. Just quickly stab it and throw the pipette
tip in LB broth+Antibiotic.

[1] http://www.level.com.tw/html/ezcatfiles/vipweb20/img/img/20297/Cell_freezing_protocol.pdf

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