DSP nuclear crosslinking

Wenduo_Ye via methods%40net.bio.net (by yewenduo from gmail.com)
Fri Apr 29 15:32:51 EST 2011


I dont know which kind of protocol are you using, but addying 1% Triton can
permeablize all the cellur fraction including Nuclear.
Did you detect if your protein of interest exist in the fraction which
supposed to be cytoplasmic?

If you prefer to use detergent to seperate your lysis , I recommend you to
use digitonin 20ug/ML and add at least 5mM Mg2+ to stablize the cell
nuclear.
Otherwise, homoginize in non detergent buffer would be much suitable for
this usage

On Thu, Apr 28, 2011 at 10:02 PM, 尹亚飞 <tuzi8780238 from gmail.com> wrote:

> I'm a student of Tsinghua University, I'm doing a crosslinking IP recently,
> and the crosslinking regent is DSP. My target protein complex is in the
> nucleus, however, during my experiment, I found my target protein of the
> crosslinked sample even could not be detected in nuclear exact, while the
> control sample(i.e., sample without crosslinking) performed well. I tried
> different lysis buffer from High Salt solution(350mM NaCl) to RIPA
> buffer(0.1%SDS, 1% Triton X-100), so I do wonder what kind of lysis buffer
> you use? Thank you!!!
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