Hello,
I am trying to find interacting protein partners for a small nuclear intracellular domain (which is my protein of interest). My plan is to do IP with nuclear extract followed by Mass spec.
Found several options in literatures but confused in choosing the proper detergent. Urea or SDS would be too harsh for maintaining weak interaction in my opinion.
Could anyone suggest me a good detergent for nuclear protein extraction that will lyse the nuclear membrane but at the same time would keep the weak interaction between proteins as well.
Thanks in advance.
Hi Tanmoyita, the most "gentle" detergent used in biochemistry is octyl glucoside.
I am sure that it has been used on nuclei.
Hi,
You can use a hypotonic buffer followd by sucrose cushion to isolate the nuclei first.
After that try using RIPA buffer with or without SDS. Use benzonase or sonication to het rid of buky DNA fragements before solubilizing your protein.
You can also perform crossliking your protein and then you dont need to care about detergent.
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