transfection quality DNA- help

John Fox J.R.Fox at shu.ac.uk
Sat Mar 18 15:19:53 EST 1995


In article <3kd9c9$o9f at no-names.nerdc.ufl.edu>, apeel at maple.circa.ufl.edu says:
>
>Am having difficulty obtaining >40% supercoiled DNA with PBPR 322 vectors using
>Qiagen columns for 500 ml cultures, even though a 1.5 sample of the same
>culture run through a Qiagen minicolumn gives marvelous supercoiling.  Don't
>have the same problem with vectors using pBS backbone.  Any ideas (the PBR
>vectors are also quite larege 9 - 11kb- is this a factor?)  Don't get much
>better results with Maniatas PEG purification and this is a much longer
>procedure.  Am not set up to do CsCl centrifugation.  Suggestions, comments,
>similar results with PBR 322 backbone vectors and these preps?

I too have had problems when I tried obtaining pGEM-3 vectors using Qiagen
minicolumns.  I resorted to CsCL centrifugation.  Sorry I can't help.

John



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