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文献和实验Thaw out 1 ml aliquot of fusion protein lysate. Centrifuge to remove precipitate. Incubate 1 ml lysate with 0.4 ml 50% GST beads. Incubate at 4C with mixing for 1 hour. Centrifuge 5K 2' in 1.5 ml eppendorf tubes. Wash the beads 4X with PBST
GST Fusion Protein Preparation
CLEAVAGEAlternate Mini PrepThaw out 1 ml aliquot of fusion protein lysate.Centrifuge to remove precipitate.Incubate 1 ml lysate with 0.4 ml 50% GST beads.Incubate at 4℃ with mixing for 1 hour.Centrifuge 5K 2' in 1.5 ml eppendorf tubes.Wash the beads 4X with PBST
Thrombin Cleavage of GST-Fusion protein
. Keep each supernatant aside 4. As a control you can elute the remaining uncleaved protein (still attached to the resin through the GST) by extraction several times with elution buffer. 5. Check results by PAGE-SDS gels or immediately separate the cleavage products
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