If you are using a reticulocyte-based system, it is chock-full of globins
that will interfere with the resolution of your protein on SDS-PAGE. This
is why you get good signal after binding. You may need to
immunoprecipitate your input protein (or separate it from the globins
some other way) in order to see it clearly.
In article <Pine.SOL.4.05.10001271548380.8853-100000 at family.sogang.ac.kr>
"Kim, Chun" <gidarim at family.sogang.ac.kr> wrote:
>> I have a problem with my GST-pulldown assay for interacting
> domain mapping. My problem is that in vitro translated input
> proteins(smaller than 25kDA) always gives me fuzzy bands.
> But I can get clear signals after binding to beads.
> I used 15% SDS PAGE.
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