Replication Origins and Two D gels... Question??
rpelle1 at po-box.mcgill.ca
Wed Jun 26 16:28:06 EST 1996
In article <31C89428.6D2A at odyssee.net>, dellaire at ODYSSEE.NET (Graham
> I am thinking of looking at the replication status of a sequence near an
> integrated vector in mouse cells. I want to know if compared to another
> locus if the region is firing (replicating) at the same time or
> differently (later or earlier... banding is not out but the two
> sequences are perhaps in the same R band).
> Can you look at origins in mammals by 2 D analysis? or do you need
> something that fires many times in a cell cycle like polyoma...etc to be
> able to see it on a 2-D gel?
> Any suggestions or comments are welcome.
> G. Dellaire
> McGill Exp. Medicine
> dellaire at odyssee.net
The 2D-gel technique has been improved enough these last years to be able
to detect single-copy origins in unsynchronized mammalian cells. I suggest
you read the papers of Joyce Hamlin and Peter Dijkwel (mostly in Mol.
Cell. Biol.) about DHFR origin mapping. In your case, you will need to
synchronize the cells, since you are interested in looking at the time of
activation. Another way is to use the PCR. Marc Lalande from Harvard has
developped such a technique.
More information about the Methods