Does ANYONE get anywhere close to 15 ug from Qiaprep Spin Columns???

I know column overloading is a potential problems, and I have tried everything from 1 - 5 ml of o/n LBA cultures. I rarely get more than 5 ug...

I would love to hear any tips you have found to get these preps to behave as promised.

Cheers,

Audra

I assume your talking about the QiaQuick columns (DNA) ? Try warming your elution buffer to 55oC and, once adding your 50ul to the column, let it incubate at 55oC for 5 or 10 minutes in a heat block. THEN spin it down. This will likely improve your yield. (it does for me). Also, try doing two elutions from the same filter column (2x 25ul or 2x 50ul), this will also help.

We routinely use the Qiagen Maxi kits, but for minis I find it works very well if you just don't use the mini columns. So just use the P1-3 solutions, spin down, transfer the SN to a fresh tube and precipitate with 0.7 volumes of Isopropanol. If I'm doing RE digests or even sequencing, I would then wash the pellet with 70% etanol, briefly air dry and resuspend in water or TE.

If you need very pure samples though, you could resuspend the pellet in about 50 L water then add about 300 L QBT (column equilibration buffer) and then load this on to a pre-equilibrated column and follow the protocol. This should allow you to reach the column's binding potential.

This pre-precipitation is also a good idea when doing maxis (or minis) on low copy number plasmids.