胚胎干细胞电转染

ElectroSquarePorator™
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

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胚胎干细胞电转染