Re: Get a plasmide - buy? share?

You can apparently get away without Lysozyme when using lab strains of
E.coli, but it *is* recommended. I suggest getting oodles of cheap
lysozyme from www.brouwland.com, where you can get it as a crystallised
powder for far, far cheaper than commercial biotech grade lysozyme. It
won't be as pure I expect, but that's not really necessary as far as
stage 1 of a miniprep is concerned. With some tweaking/trial/error you
should be able to find an amount that works just as well for less than a
hundredth of the price.

I'd stick with sodium salts of EDTA, but it might be worth getting the
lower sodium contents as you can use this more easily with DIY
electrophoresis buffers, too. The more sodium in your EP buffers, the
more current and the more heat. In the original paper on using Sodium
Borate, the authors dedicated part of their rant-against-TAE/TBE to the
sodium content of the dissolved EDTA.

On 28/12/11 15:34, Ethan wrote:
> Mega, While I have never done the prep, I would not recommend skipping
> the lysozyme. From my understanding, its purpose is to break down
> parts of the bacterial cell wall (namely peptidoglycan). I imagine
> that eliminating it from the procedure would cause either a drastic
> drop in yield or total lack of yield.
>
> I am looking into several reagents, and I was wondering if there is
> any functional difference between the various salts of EDTA available.
> My guess would be that the disodium and tetrasodium salts would only
> differ in the dissolved pH, and so might need additional adjusting
> accordingly. However, having a calcium salt of EDTA seems
> counterintuitive to me because it functions to chelate several ions in
> solution, including Ca2+. Which salt is typically used in the
> laboratory? Thanks!
>
> On Dec 28, 7:14 am, Mega <masterstorm...@gmail.com> wrote:
>> So my shoping list for plasmid miniprep (boiling lysis):
>>
>> STET-Buffer
>> lysozyme (really necessary??)
>> water
>> isopropanol

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