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Chris:LabNotes/General Protocols/Electroporation
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==Protocol== *'''On ice,''' we want to add 50-60ul of cells + small volume of DNA plasmids (max 5ul, fg levels) *Turn on machine and set volts to 1800 (based on specification of cuvettes) *Take off the cap of the cuvette and discard. Pipette in samples into the middle of metal plates/slot within cuvette (will not be able to reach all the way down with pipette) *Tap the cuvette to make sure cells go to the bottom in between the two metal plates/slot and keep on ice *Take cuvette out of ice and wipe with kimwipe to remove precipitation on sides *Put cuvette in plastic holder (will have a notch on one side of cuvette so there is only one way to place into holder) *Push holder into slot in machine such that the metal sides of the cuvette make contact with metal plates in machine *Press "Pulse" twice and wait for beep *'''Immediately''' take out cuvette from holder and add 250ul SOC and pipette up and down. It is essential to do this immediately after pulse/beep in order to save the bacteria *Use p200 pipette to transfer as much of the cells from cuvette into an Eppendorf tube (can tilt cuvette slightly to optimize volume of cells recovered) *Incubate at 37C for a few hours and plate or grow in liquid media with antibiotic overnight *Discard cuvette after use
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