precipitating from Qiaquick

Gene Huh gshuh at garnet.berkeley.edu
Sun Sep 22 06:39:16 EST 1996


In article <51u40m$hp9 at paperboy.ccc.nottingham.ac.uk>,
pdxrmb at vaxb.ccc.nottingham.ac.uk wrote:

# I have been using Qiagens qiaquick columns for the purification of PCR
products and bands form gels. Direct elution yields good quality DNA, but
the concentration is a little low - I would prefer a nice high
concentration stock for storage etc. Ethanol precipitation produces very
low yields (as judged from EtBr gels)despite there being plenty of DNA in
the eluate- is there a reason for this ?
# I thought maybe carryover of chaotropic salt may effect the
precipitation conditions ? but wasnt exactly ur why this shoud be. any
advice/infromation would be gratefully received. thanks in advance-
Richard M Badge.

I have found this to be the case as well -- that is, no more than half of
the expected yield when agarose gel-purified fragments are carried thru
Qiaquick columns.  I didn't add glycogen or any other carrier, since I
often see a nice pellet (agarose?).  One possibility might be that the pH
of the eluate (8.5 or thereabouts) is nonoptimal for precipitation. 
Another is that the aforementioned pellet may contain mostly non-LMP
agarose -- perhaps that's a lot harder to resuspend?  I guess the latter
explanation wouldn't apply for PCR cleanup, tho'.  With respect to
chaotropic salts,  I recall that RNA pellets from acid phenol/guanidinium
thiocyanate procedures are often quite recalcitrant to resuspension,
although in this latter case nucleic acids are precipitated from a very
high concentration of GTC.  

Hope this helps?
Gene Huh
gshuh at garnet.berkeley.edu



More information about the Methods mailing list