De-crosslinking DNA-protein IPs

EK khatipovNO-SPAM at NO-SPAMuchicago.edu
Wed Oct 23 15:39:19 EST 2002



Hi everybody,
A colleague of mine is struggling with the following question. Would boiling
in standard SDS-PAGE loading buffer de-crosslink the proteins she fished our
via IP with the Abs against one of the proteins in the crosslinked complex?
She used a DNA-binding protein (nuclear) as a bait. Before IP, she
crosslinked DNA and proteins with paraformaldehyde. There is a protocol she
knows that says incubating at 65C overnight with 1%SDS and high salt will
reverse crosslinking, but I am quite skeptical about that. Is this at all
possible to reverse PFA crosslinking ?
-Emir






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