DNA prep for transient transfection

seppen at amc.uva.nl seppen at amc.uva.nl
Mon Mar 6 12:49:53 EST 1995

In Article <nick.5.2F559DCF at lmb1.rug.ac.be>
nick at lmb1.rug.ac.be writes:
>Hi there,
>As a coincidence, we had a discussion only yesterday about problems with low 
>efficiencies of transient transfections in our lab.
>More precisely, transfections using the DEAE-dextrane method seem to give very 
>variable results, given one plasmid preparation to another.
>The low efficiency is independent of the person, the acceptor cells or the DNA 
>itself. In fact...the problem seems to be the preparation method used. Sorry 
>to tell you so, but this all began when we started using QIAGEN columns!
>I am not trying to start a "WIZARD (TM)"-like hystery here. We just want to 
>know if other people have experienced similar problems, and if there is any 
>solution to this. Especially if you want to compare two different genes, this 
>varying efficiencies really make it a mess.
>I hope someone has an answer to this problem.
>	Greetings,
>	Nick
>Laboratory of Molecular Biology
>University of Gent
>Nick at lmb1.rug.ac.be

Hi nick,

we had a similar problem with obtaining stable transfectants. I now routinely
do one phenol and two chloroform/iaa exctractions after the maxiprep with
qiagen. The interfase always contains protein, so qiagen is probably not
enough to get pure vector.

Jurgen Seppen
The Netherlands

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