Protein Expression and Purification Series - Bio-Rad

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CHAPTER 8 BIOLOGIC DUOFLOW PROTOCOL Protein Expression and Purification Series Concentration of GST-DHFR-His (Desalted fraction A) = ______________________ M Concentration of GST-DHFR-His (Desalted fraction B) = ______________________ M Concentration of GST-DHFR-His (Desalted fraction C) = ______________________ M B: Convert from molarity to mg/ml for the amount of GST-DHFR-His in your purified desalted fractions knowing that the molecular mass of GST-DHFR-His is 50,361 g/mol. Concentration GST-DHFR-His (mg/ml) = Concentration GST-DHFR-His (M) x 50,361 g/mol Concentration of GST-DHFR-His (Desalted fraction A) = _________________________mg/ml Concentration of GST-DHFR-His (Desalted fraction B) = _________________________mg/ml Concentration of GST-DHFR-His (Desalted fraction C) = _________________________mg/ml These are the concentrations of GST-DHFR-His that you produced and purified and are contained in your 150 µl desalted fractions. 192 Chapter 8: Purification Protocol for BioLogic DuoFlow System
Protein Expression and Purification Series SDS-PAGE Electrophoresis Analysis of Purified Fractions Student Workstations Each student team requires the following items to analyze their purification samples via SDS-PAGE: Material Needed for Each Workstation Quantity Soluble PAGE sample 100 µl Flowthrough PAGE sample 100 µl Wash PAGE sample 100 µl Fraction A PAGE sample 100 µl Desalted A PAGE sample 50 µl Fraction B PAGE sample 100 µl Desalted B PAGE sample 50 µl Fraction C PAGE sample 100 µl Desalted C PAGE sample 50 µl Precision Plus Protein Dual Color standards 15 µl 4–20% Mini-PROTEAN TGX precast gel 1 Vertical electrophoresis chamber (per 1–4 gels) 1 1x Tris/Glycine/SDS (TGS) running buffer (per electrophoresis chamber) 700–1,200 ml Power supply (200 V constant) to be shared between workstations 1 Buffer dam (only required if running 1 gel/box) 1 2–20 µl adjustable-volume micropipet and tips 1 Staining tray 1 Bio-Safe Coomassie stain (per gel) 50 ml Marking pen 1 Common Workstation Quantity Water bath or heat block set to 95°C 1 Microcentrifuge with variable speed setting >16,000 x g 1 Water for gel washing and destaining (tap water is fine) 1 L Chapter 8: Purification Protocol for BioLogic DuoFlow System 193 CHAPTER 8 BIOLOGIC DUOFLOW PROTOCOL
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Biotechnology Explorer TM Protein E
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Protein Expression and Purification
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CHAPTER 1 BACKGROUND Protein Expres
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CHAPTER 2 FOCUS QUESTIONS Protein E
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CHAPTER 3A ADVANCE PREP CENTRIFUGAT
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CHAPTER 3B ADVANCE PREP INSTRUMENTA
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CHAPTER 4 11 ml CULTURE PROTOCOL Pr
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CHAPTER 5 100 ml CULTURE PROTOCOL P
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CHAPTER 6 HANDPACKING COLUMNs Prote
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CHAPTER 7 BIOLOGIC LP SYSTEM PROTOC
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Page 218 and 219: APPENDIX C RCF TO RPM CONVERSTION P
Page 220 and 221: APPENDIX D SETTING UP THE SMARTSPEC
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Page 224 and 225: APPENDIX E MINI-PROTEAN GEL BOX SET
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APPENDIX I GLOSSARY Protein Express
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APPENDIX I GLOSSARY Protein Express
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APPENDIX J REFERENCES, LEGAL NOTES
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Life Science Group Bulletin 1665067
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Protein Expression and Purification Series - Bio-Rad

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