Is ChIP is a good choice to find the DNA sequence for a binding protein? - (Oct/24/2006 )

Hi !

I want to find the DNA sequence that my protein binds too. I know that my protein is found in nucleus and have a putative nuclear receptor site.

Is ChIP is a good method to find the DNA sequence that can bind to my protein ? I suppose that after the immnunoprecipitation I have the amplified the unknown region that was crosslinked. Is there any random primer that can I use for PCR ? Is it any other method that can be use to find the DNA sequence of a specific DNA binding protein ?

thanks for your help

Hi Christian,

ChIP is a method to work out the DNA sequence your protein is binding to, you would clone the IP DNA and sequence many to see what has been pulled down, as ChIP is a "noisy" assay in terms of binding (you get non-specific IP of some fragments) so the more clones you sequence the higher the chance of actually picking up the real sequence your protein is binding to.

You could go the "old technology" of DNA footprinting but with this, you would need to have a fair idea what your sequence would be. have you got a narrow list of candidate sequences to choose from?

N

Some people are doing ChIP on chip that means they do a ChIP assay and then they hybridize a chip (array) to check the IPed sequences. I haven't done it myself but I guess that would be the "fastest" way...