Protein Expression and Purification Series - Bio-Rad

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Protein Expression and Purification Series Materials Required But Not Supplied Chemicals Centrifugation BioLogic LP/BioLogic Purification DuoFlow Purification Dry ice 1 block 1 block Ethanol 0.5–2 L 0.5–2 L Plastics and Consumables Centrifugation BioLogic LP/BioLogic Purification DuoFlow Purification 100–1,000 µl pipet tips, standard style 12 boxes 4 boxes (catalog #223-9350EDU) 2–200 µl pipet tips, standard style (catalog #223-9347EDU) 12 boxes 4 boxes Parafilm sealing film 1 1 UV compatible cuvettes such as trUView cuvettes (catalog #170-2510EDU) 1 box of 50 1 box of 50 or or or quartz submicrovolume cuvettes (catalog #170-2505EDU) 1–4 cuvettes 1–4 cuvettes 1.5 ml standard disposable polystyrene cuvettes (catalog #223-9955EDU) 1 box of 100 1 box of 100 or or quartz standard cuvettes (catalog #170-2502EDU) 1–4 cuvettes 1–4 cuvettes 10 ml syringes 12 16 22 gauge syringe needles 12 4 Fraction collection tubes (catalog #223-9751EDU) N/A 200 tubes Glassware Centrifugation BioLogic LP/BioLogic Purification DuoFlow Purification 500 ml Erlenmeyer flasks N/A 4 1 L Erlenmeyer flask or autoclavable bottle 2 1 Beakers for dry ice/ethanol bath 1–12 1–4 Temperature Control Equipment Centrifugation BioLogic LP/BioLogic and Mixing Devices Purification DuoFlow Purification Microwave oven 1 1 –20°C freezer 1 1 Incubation oven (catalog #166-0501EDU) 1–2* 1 Tube roller (catalog #166-0711EDU) 1–2* N/A Mini rocker (catalog #166-0710EDU) 1** 1 Shaking incubator or shaking water bath 1* N/A capable of holding 4 x 500 ml flasks N/A 1 Dry bath (catalog #166-0562EDU) or water bath 1 1 * Initial overnight cell cultures and cell subcultures/induced cells for Centrifugation Purification are prepared in 50 ml sterile conical tubes. Incubation at 37°C with some form of mixing is required. This incubation can be accomplished in either a temperature-controlled shaking incubator, a shaking water bath or using tube rollers in an incubation oven set at 37°C. The Bio-Rad tube roller can hold six 50 ml sterile conical tubes. Therefore, if 12 workstations are being run, two tube rollers, each in their own incubation oven, would be required. ** For the Centrifugation Purification protocol, during the binding of the cell lysate to the Profinity IMAC Ni-charged resin, a tube roller can be used for end over end mixing of the resin and lysate. If a tube roller is not available, the mixing can be accomplished by using a mini rocker. The mini rocker can be further used for rinsing, staining and destaining of SDS-PAGE gels. 12 Materials Required But Not Supplied
Protein Expression and Purification Series Other Equipment Centrifugation BioLogic LP/BioLogic Purification DuoFlow Purification UV/Vis Spectrophotometer capable of reading 1 1 to three decimal places (catalog #170-2525EDU) 100–1,000 µl adjustable micropipet (catalog #166-0508EDU, 166-0553EDU) 12 4 20–200 µl adjustable micropipet 12 4 (catalog #166-0507EDU, 166-0552EDU) 2–20 µl adjustable micropipet 12 4 (catalog #166-0506EDU, 166-0551EDU) Pipet pump or filler 12 4 10 ml serological pipets 4 12 Power supply (catalog #164-5050EDU) 3–12 1–4 Vertical electrophoresis chambers 3–12 1–4 (catalog #165-8004EDU, 165-8005EDU) Gel documentation system 1 1 (catalog #170-8170EDU, 170-8270EDU) Microcentrifuge with variable speed setting 1 1 capable of 16,000 x g (catalog #166-0602EDU) Centrifuge capable of 16,000 x g N/A 1 with rotors that hold 250 ml centrifuge bottles and 30–50 ml centrifuge tubes Centrifuge bottles (250 ml) capable of withstanding 4,500 x g N/A 4 Centrifuge tubes (30–50 ml) N/A 4 capable of withstanding 16,000 x g Handpacked Column Process Only Quantity Flow adaptor, 1.0 cm column ID 1 per instrument 1–7 cm functional length (catalog #738-0014EDU) Equipment Necessary If Using a BioLogic DuoFlow System (catalog# 760-0047EDU or 760-0037EDU) Quantity Fittings to convert luer to ¼-28 (catalog #732-0113EDU) 1 set per instrument 5 ml injection loop (catalog #750-0497EDU) 1 per instrument Miscellaneous Centrifugation BioLogic LP/BioLogic Purification DuoFlow Purification Marking pens 12 4 Storage boxes for microcentrifuge tubes 3 1 (catalog #166-0482EDU) Green racks for microcentrifuge tubes 12 4 (catalog #166-0481EDU) Materials Required by not Supplied 13
Page 1: Biotechnology Explorer TM Protein E
Page 4 and 5: Protein Expression and Purification
Page 34 and 35: CHAPTER 1 BACKGROUND Protein Expres
Page 50 and 51: CHAPTER 2 FOCUS QUESTIONS Protein E
Page 56 and 57: CHAPTER 3A ADVANCE PREP CENTRIFUGAT
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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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CHAPTER 8 BIOLOGIC DUOFLOW PROTOCOL
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CHAPTER 9 DHFR ENZYMATIC ASSAY Prot
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APPENDIX A TIPS AND HELPFUL HINTS P
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APPENDIX A TIPS AND HELPFUL HINTS P
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APPENDIX B RESULTS ANALYSIS Protein
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APPENDIX B RESULTS ANALYSIS Protein
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APPENDIX C RCF TO RPM CONVERSTION P
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APPENDIX D SETTING UP THE SMARTSPEC
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APPENDIX D SETTING UP THE SMARTSPEC
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APPENDIX E MINI-PROTEAN GEL BOX SET
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APPENDIX F BIOLOGIC LP SYSTEM SETUP
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APPENDIX F BIOLOGIC LP SYSTEM SETUP
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APPENDIX G BIOMANUFACTURING Protein
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APPENDIX G BIOMANUFACTURING Protein
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APPENDIX H FOCUS QUESTIONS INSTRUCT
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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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