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isolation of protein complex but not with BN PAGE

isolation of protein complex but not with BN PAGE - Protein Forum

isolation of protein complex but not with BN PAGE - Protein Forum


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Old 12-13-2010, 06:46 PM
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Default isolation of protein complex but not with BN PAGE



Dear all,
I need some suggestions, I'm trying to isolate a complex protein (a transmembrane protein complex which bind to a receptor) from plasma membrane of arabidopsis. I used pull-down assay for isolating the complex by using a column for bind strep tag which constructed in C terminus of receptor protein. Moreover, I apply 1% digitonin as the detergent for releasing the complex from plasma membrane. Unfortunately, so far no a complex protein that I can isolate. Only the fractions of the receptor protein and other unknow proteins which are shown up in the native gel.
Some friends told me to use BN PAGE for isolating the complex protein, since BN PAGE can maintain native condition of the complex. I tried once but it seems that it is not suitable for my complex protein. The protein that i want to isolate, according publication, is predicted very rare (only a little amount in the cells) thus now i'm really stuck with my work.
Perhaps be there is someone who has experiences in isolating of native protein, please give me some advices and suggestions.
any suggestion and advices will be very helpful for me

thank's

best regard
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