cytoplasm and nuclear run off - cytoplasm and nuclear run off (Jan/13/2011 )

I want to seperate nucleus from cytoplasm and pool the cytoplasm, transfect a plasmid to cytoplasm and then add nucleus to demonstrate transcription. Has anyone done a simialr work and if so how did you go about seperating and pooling the nucleus and cytoplasm

Thank you,

Herbie

So basically doing a sub-cellular fractionation and then adding nuclei? You are aware that this will destroy the cells unless you are planning on removing the nuclei individually from the cells by patch-clamping style techniques?

bob1 on Thu Jan 13 22:23:37 2011 said:

You can isolate intact nuclei by sub-cellular fractionation. If you want viable cells out the end, you will need to investigate techniques that remove the nucleus by micromanipulation (look for protocols around cloning of animals - most people do this with animal eggs).