Reporter protein to test NLS function

jlight at SICKKIDS.ON.CA jlight at SICKKIDS.ON.CA
Sun Jun 7 20:14:36 EST 1998


I need a good reporter protein to test the function of NLS motifs in
mammalian cells i.e. I want to fuse the NLS to the reporter protein and
show that the fusion protein goes into the nucleus if the NLS is
functional.  Ideally, the reporter proteins should be inert, stable, bigger
that 60 kD, strictly cytoplasmic but capable of nuclear localization.  I've
tried using lacZ, probing it with an anti-beta-gal antibody, but I
generally see signal in both the cytoplasm and nucleus with vector
transfections only (no signal with non-transfected cells though).  I'm
assuming that beta-gal isn't that stable when expressed in mammalian cells,
and that I'm picking up parts of beta-gal that have diffused into the
nucleus.  I would really like to hear from somebody who has done or is
doing these kind of experiments, and has a good system.  Thanks in advance
for your help.






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