purification:tev:c_protocol
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| purification:tev:c_protocol [2016/07/18 08:32] – bonnefon | purification:tev:c_protocol [2016/07/18 08:33] (current) – bonnefon | ||
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| * The sample was then applied to 1 mL of Ni-NTA agarose (GE Healthcare) pre-equilibrated with Buffer A((25 mM sodium phosphate (pH 8.0), 10% (v/v) glycerol, 200 mM NaCl, and 25 mM imidazole)). | * The sample was then applied to 1 mL of Ni-NTA agarose (GE Healthcare) pre-equilibrated with Buffer A((25 mM sodium phosphate (pH 8.0), 10% (v/v) glycerol, 200 mM NaCl, and 25 mM imidazole)). | ||
| * The protein was eluted with Buffer B((25 mM sodium phosphate (pH 8.0), 10% (v/v) glycerol, 200 mM NaCl, and 500 mM imidazole)). | * The protein was eluted with Buffer B((25 mM sodium phosphate (pH 8.0), 10% (v/v) glycerol, 200 mM NaCl, and 500 mM imidazole)). | ||
| - | * The eluted protein was then applied to a Superdex 75 (16/10) column in Buffer C((25 mM sodium phosphate (pH 8.0), 10% (v/v) glycerol, 200 mM NaCl, 5 mM β-mercaptoethanol)). | + | * The eluted protein was then applied to a Superdex 75 (16/10) column in Buffer C((25 mM sodium phosphate (pH 8.0), 10% (v/v) glycerol, 200 mM NaCl, and 5 mM β-mercaptoethanol)). |
| * The eluted fractions containing TEV were pooled, flash-frozen in liquid nitrogen, and stored at −80°C. | * The eluted fractions containing TEV were pooled, flash-frozen in liquid nitrogen, and stored at −80°C. | ||
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