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T820 ELECTROPORATION PROTOCOL Mouse Embryonic Stem (ES Cells) Cell Preparation: Growth Medium: DMEM + 10%HS+Non Essential Amino Acids+B-ME+LiF Trypsinization: 0.5ml Trypsin-EDTA per 60mm dish for 5 minutes at 37°C. Stop with 1ml media Washing Procedure: Pellet by centrifugation. Decant and resuspend in HBS. Repeat wash Cell Density: 107 cells/ml Electroporation Settings: Choose Mode: LV Set Voltage: 500V Set Pulse Length: 99μs Set Number of Pulses: 1 Chamber: BTX Disposable Cuvette P/N 620 (2mm gap) Desired Field Strength: 2.5 kV/cm Electroporation Procedure: Sample Volume: 400μl DNA Concentration: 40-80μg/ml Temperature: Room Temperature Pulse: Press the Start button Incubate cells for 10 minutes Plate: Dilute with 5ml of media and plate on 60mm petri-dish Results: Positive transfection Reference: Personal Communication, Dr. Jonathon Dinsmore, Diacrin Inc., Charlestown, MA JIMMY XU提供 |
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Culturing HEK 