episomal vectors

Ian A. York iayork at panix.com
Mon Aug 25 09:20:28 EST 1997


In article <34026CA5.6EDE56F0 at leland.stanford.edu>,
Wensheng Chen  <wensheng at leland.stanford.edu> wrote:
>I am looking for an episomal vector that can replicate in mouse cell
>lines but does require a prior transfection of cells with genes such as
>SV40 large T antigen. The goal is to do expression cloning with
>libraries made in such vector. Unfortunately the literature has been

Do you mean "does not" require transfection with SV40 LT?  (SV40 LT
wouldn't help in mouse cells anyway, because SV40 only works in primate
cells, basically.)  There are mouse cells transformed with polyoma
large-T antigen, though, which drive replication of plasmids containing
the polyoma virus origin of replication;  PyOri is present in many of
Invitrogen's vectors (the ones derived from pCDM8), which are really
library vectors.  These vectors don't replicate to as high levels as do
SV40-ori plasmids in SV40-LT cells, but they do work in transients.  

Ashok mentions EBV/chromosomal origin vectors that replicate in mouse
cells.  I'm not sure that these would be well-suited for library cloning,
because (as I understand it, which might easily be wrong) these are fairly
large vectors and you may have trouble getting representative and
appropriately-sized inserts in them.

If the PyOri vectors are unsuitable, for some reason, then I think you
have a problem and may need to consider going with stable transfection of
the library--you can still do expression cloning this way, though it's
trickier than by transient transfection.

Ian
-- 
      Ian York   (iayork at panix.com)  <http://www.panix.com/~iayork/>
      "-but as he was a York, I am rather inclined to suppose him a
       very respectable Man." -Jane Austen, The History of England



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