alternatives to southern or FISH?
hansw at sci.kun.nl
Sun Aug 25 07:38:39 EST 2002
Peter Ianakiev <ianakiev at genome.wi.mit.edu> wrote in message news:<3D57E410.B4DE3644 at genome.wi.mit.edu>...
> Volker Budinger wrote:
> > Hi,
> > I am characterising some cell lines which were transfected with a
> > linearised plasmid. Now i'm searching for a way to find out, how often
> > this single sequence was inserted into the genome of those HeLa's.
> > Probably this could be done by southern or FISH but I'am searching for
> > other alternative methods instead of these. (Cause I've got problems
> > with the sensitivity of the southern assay and fish could not be done in
> > our laboratory.)
> > Does anyone know alternative methods?
> > Thanks
> > Volker Budinger
> > ---
Try to perform REAL-TIME PCR... works great with cell-lines and it is
possible to estimate the relative percentage of your sequence in
comparison to a house-hold gene (reference gene, or as you might even
say blanco ;-) )
The only problem with this is that the real-time pcr machiene costs
very very VERY expensive... But if you know an other department who
has this machiene and knowledge then you can have beautiful
It is often used in this kind of problems offering great sensetivity
and relative comparison.
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