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protocol to keep cytoplasmic lysates


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#1 labstud4

labstud4

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Posted 15 August 2013 - 08:29 PM

Does anyone have a ChIP protocol that will allow me to perform IP on the nuclear fraction with chromatin, while keeping the cytoplasmic proteins so I can run a traditional IP on a Western? I would like to be able to IP the nucleus and do PCR and also IP the cytoplasm and run a Western.

 

Many protocols call for a cell lysis buffer to pellet the nuclei; can I just simply keep this sup for a Western? They also use 10ml of the cell lysis buffer to pellet the nuclei; can I get away with using less in order to concentrate the proteins?

 

Alternatively, can I perform regular ChIP up until the IP portion, but then add DNase to get rid of the chromatin? Ideally, I would like to isolate the nuclear transcription factors and perform an IP then a Western to see some co-factors.

 

Edit: Please delete this topic.


Edited by labstud4, 15 August 2013 - 08:48 PM.





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