Sauer:Purification of His-tagged proteins/Denaturing prep: Difference between revisions
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m (Sauer:Purification of His-tagged proteins/Deanturing prep moved to Sauer:Purification of His-tagged proteins/Denaturing prep) |
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10 mM imidazole | 10 mM imidazole | ||
Adjust to pH 8 | |||
===Wash buffer 1=== | |||
100 mM NaH<sub>2</sub>PO<sub>4</sub> | |||
150 mM NaCl | |||
8 M urea | |||
20 mM imidazole | |||
Adjust to pH 8 | |||
===Wash buffer 2=== | |||
50 mM NaH<sub>2</sub>PO<sub>4</sub> | |||
500 mM NaCl | |||
20 mM imidazole | |||
Adjust to pH 8 | |||
===Elution buffer=== | |||
50 mM NaH<sub>2</sub>PO<sub>4</sub> | |||
500 mM NaCl | |||
250 mM imidazole | |||
Adjust to pH 8 | Adjust to pH 8 |
Revision as of 14:49, 8 February 2006
This is a general protocol for purification of His6-tagged proteins under denaturing conditions. Note that you may need to modify this protocol to fit your specific needs.
Buffers
Lysis buffer
100 mM NaH2PO4
10 mM Tris base
6 M guanidine hydrochloride
10 mM imidazole
Adjust to pH 8
Wash buffer 1
100 mM NaH2PO4
150 mM NaCl
8 M urea
20 mM imidazole
Adjust to pH 8
Wash buffer 2
50 mM NaH2PO4
500 mM NaCl
20 mM imidazole
Adjust to pH 8
Elution buffer
50 mM NaH2PO4
500 mM NaCl
250 mM imidazole
Adjust to pH 8