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kx444555: ½ð±Ò+2, ¹ÄÀø½»Á÷ 2015-04-09 08:24:17
Inclusion bodies were separated from the cytosolic protein pool by centrifugation at 20,000 g for 30 min, and the inclusion bodies-containing pellet was solubilized with 4 ml of solubilization buffer (6 M Urea, 200 mM NaCl, 100 mM Tris-HCl, pH 8.3), supplemented with 10 ¦ÌM ¦Â-mercaptoethanol and incubated overnight at 4¡ãC under constant rotation. Thereafter, the sample was spun at 25,000 g for 30 min to remove any insoluble material and the His-tagged protein was purified using HisPurTM Ni-NTA Resin

Hope it works with you. Good luck!
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2Â¥: Originally posted by Ä«¶û±¾»úÆ÷ at 2015-04-08 19:37:55
Inclusion bodies were separated from the cytosolic protein pool by centrifugation at 20,000 g for 30 min, and the inclusion bodies-containing pellet was solubilized with 4 ml of solubilization buffer ...

thank you for your attention
Some details need to be confirmed£º
1,Urea is a denaturant, whether the protein is denatured after dissolution
2,The need for  agitation when constant rotated£¿
3,the sample was spun at 25,000 g for 30 min to remove any insoluble material ,so The supernatant is  the soluble protein£¬right£¿
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