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specific phosphorylation

kinase phosphorylation

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2 replies to this topic

#1 simone dorfmueller

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Posted 19 February 2013 - 07:47 PM

Hi.
I am interested in the target of one specific kinase.
So I was thinking of dephosphorylating cell lystae (which shouldn't be a problem) and than to incubate the dephosphorylated lysate with my kinase and see what is re-phosphorylated.
of course this means that my proteins must be functional after dephophorylation (so no boiling, dtt etc), and ideally, but probably nearly impossible, to supress all kinases except the one I am adding.

Did anybody ever did this? or heard of somebody doing this?

Cheers
Simone

#2 bob1

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Posted 20 February 2013 - 12:09 AM

How about in-vitro translation using rabbit reticulocyte lysate?

#3 Joe KooL

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Posted 11 November 2013 - 09:02 PM

HI SImone,

I've recently purchased lambda phosphatase and am wondering how to dephosphorylate cell lysates. Do you mind sharing your protocol?







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