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DNA re-suspension

DNA re-suspension - DNA Forum

DNA re-suspension - Discuss DNA, the molecule of hereditary. Topics include DNA structure, DNA replication, DNA function.


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Old 04-19-2010, 03:11 PM
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Default DNA re-suspension



Hello,

First time user.

I have been conducting an experiment where I extract plasmids from bacterial cells.

I was wondering what is a good re-suspension buffer for DNA other than EB buffer? The reason is for storage pursposes.

any help would be greatly appreciated!
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Old 04-19-2010, 03:37 PM
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Default Re: DNA re-suspension

TE (tris-edta), pbs, molecular grade water.
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Old 04-19-2010, 04:03 PM
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Default Re: DNA re-suspension

thanks danfive

I haven't heard of the molecular grade water. Could you direct me on where I can get more info on molecular grade water.

Thanks
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Old 04-19-2010, 04:47 PM
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Post Re: DNA re-suspension

Quote:
Originally Posted by boson8 View Post
thanks danfive

I haven't heard of the molecular grade water. Could you direct me on where I can get more info on molecular grade water.

Thanks
MG water can be chromasolv(brand name) hplc water (from Sigma) or autoclaved MilliQ water (from lab MilliQ system).
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Old 04-20-2010, 03:31 PM
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Default Re: DNA re-suspension

thanks again for the info.

I was also wondering what other buffer are commonly used to dissolve DNA (plasmids) in other than Buffer EB which is what I am using currently.

EDTA is something I want to avoid since it is known to inhibit downstream enzymatic or sequencing reactions which is essential for my experimental analysis.

angain any advice would be really helpful.
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Old 04-21-2010, 12:43 PM
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Post Re: DNA re-suspension

Quote:
Originally Posted by boson8 View Post
thanks again for the info.

I was also wondering what other buffer are commonly used to dissolve DNA (plasmids) in other than Buffer EB which is what I am using currently.

EDTA is something I want to avoid since it is known to inhibit downstream enzymatic or sequencing reactions which is essential for my experimental analysis.

angain any advice would be really helpful.
Under Qiagen FAQ page you can see that they commonly use TE and STE. You can just leave out the EDTA.

Tris (TE without the EDTA).
Also STE (100mM NaCl, 10mM Tris-Cl, 0.1mM EDTA) leave out the EDTA as desired.

BTW, if you can keep the low 0.1mM EDTA it will benefit your storage, as it binds metal cofactors (Mg++) which reduces/stops a lot of enzyme activity, like DNAse (which can come from the cell prep, your skin, lab).

The low EDTA in the small volume of storage buffer, is usually diluted out and won't affect typical PCR/sequencing reaction/restrictions digests.

Hell you can even use spin columns (GE-healthcare G25 microspin columns) to buffer exchange the EDTA out before your downstream application (its about 5-8minute procedure).
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