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Protein purification from Inclusion Bodies [View Printable]
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Pinal Pandya
Group: Member Posts: 12 Joined: Apr 24, 2006
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Hi there, I am trying to solubilize my protein from Inclusion bodies. I used DH5 alpha strain for the expression. And i found this protocol online for protein refolding. It says i need to dialyse the protein using urea. But my problem is i dont know where to buy the dialysis bags from. I am stumped !!! Can anyone help me. Thanks Pinal
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| Posted Oct 12, 2007, 19:17 PM |
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R Bishop
Group: Moderators Posts: 249 Joined: Jan 17, 2006
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Pinal, I find that Pierce has some nice dialysis products. I use the Snakeskin for large amounts of solution and the slide-a-lyzer cassettes for smaller volumes. http://www.piercenet.com/products/browse.cfm?fldID=040101Also trying using the Solution Search link on this page as there are many threads about inclusion bodies and refolding Hope that helps Rb
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| Posted Oct 12, 2007, 21:33 PM |
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Pinal Pandya
Group: Member Posts: 12 Joined: Apr 24, 2006
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Thank you soo much for your help RB , this definitely helps a lot and i am going to check out the solutions search link.
thanks again
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| Posted Oct 12, 2007, 21:41 PM |
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Pinal Pandya
Group: Member Posts: 12 Joined: Apr 24, 2006
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Thanks for helping me before RB. I have another question, I found this protocol online for solubilization and refolding. the method is as follows: add 100mM tris buffer ph 8, 50mM glycine ; sonicate; dissolve supernatant in the above buffer + 8M urea.
then add 5mM reduced glutathione and 0.5 mM oxidized glutathione. and then proceed for dialysis.
so i guess my question is why do we need ot add glutathione at all? reduced and oxidized? and if we use oxidized glutathione with ~6% ethanol , is it ok?
Please help me!!! Pinal
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| Posted Oct 16, 2007, 17:43 PM |
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