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Avoiding non-specific DNA binding in ChIP assays - (Oct/19/2005 )

Hi again, I have another doubt related to CHiP assays. I have added BSA and sonicated DNA to the protein A Sepharose, but I keen op founding in controls taht DNA (in a strain without tagging) binds to the slurry...How can I block the resin? sad.gif

-Research will kill me....-

You should also try blocking with sheered salmon sperm DNA (you can buy this cheap). In addition check your protocol for your wash conditions, it sounds like your washes may not be stringent enough to remove the non-specific binding going on.

-tap14-

QUOTE (Research will kill me.... @ Oct 19 2005, 11:51 AM)
Hi again, I have another doubt related to CHiP assays. I have added BSA and sonicated DNA to the protein A Sepharose, but I keen op founding in controls taht DNA (in a strain without tagging) binds to the slurry...How can I block the resin? sad.gif


I use sperm DNA + BSA for blocking, and I wash using Staph-A-Buffer + sodium deoxycholate.

-Research will kill me....-

Does anyone know how much salmon sperm to add to beads incubated with antibody?
In the past I have only blocked with BSA...

-beccamitsch-