Hello,
The cytoplasm often contains many membrane bound compartments which
could exclude a water soluble dye. The nucleus has a much smaller volume
fraction inaccessible to such a dye. Therefore the nucleus may have an
apparently greater concentration of the dye assuming the dye freely enters
the nucleus. Perhaps if you fuse GFP to something so large that it can't
enter the nucleus by diffusion but can only enter as a result of the nuclear
targetting sequence you will acheive the desired result.
Cheers,
Jerry
Gordon at med.unc.edu
Ettensohn Lab wrote:
> Hello!
>> I am working on a GFP fusion protein which localizes to the nucleus in
> sea urchin embryos. I have tried a control experiment in which I
> microinject only GFP - and this localizes throughout each cell
> (cytoplasm, membrane, nucleus) but concentrates in the nucleus. Does
> GFP alone normally accumulate in the nucleus? Can I be sure that the
> localization I see with my fusion protein is entirely due to the
> localization signal of protein and not GFP? Can anyone give me a
> reference in which to find out if this is a common phenomenon? Finally,
>> does anyone know of a fluorescent molecule or dye that diffuses evenly
> thoroughout a cell and does not get partitioned into the nucleus or
> other subcellular structures?
>> Thank you so much,
> Heather
