DNA re-suspension

[boson8]

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!

[danfive]

TE (tris-edta), pbs, molecular grade water.

[boson8]

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

[danfive]

Originally Posted by boson8

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).

[boson8]

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.

[danfive]

Originally Posted by boson8

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).