Keating:Transformation

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Transformation

 * written by Christy

Transformation protocol (for pure plasmid)


 * 1) Do in 1.5mL eppendorf tubes
 * 2) 50-100ul competent cells (XL1-Blue or BL-21 that were thawed on ice) + 2ul DNA
 * 3) Mix by inverting
 * 4) Place on ice for 10 minutes
 * 5) Place in heat block at 42 degrees C for 2 minutes
 * 6) Place on ice for 2 minutes
 * 7) Add 700ul of LB
 * 8) Incubate at 37 degrees C for 40 minutes
 * 9) Plate about 100ul of cells on antibiotic plate
 * 10) Incubate at 37 degrees C overnight

Transformation after ligation reactions


 * 1) Use ~150ul compentent XL1-Blue
 * 2) Place on ice for 10 minutes
 * 3) Place at 42 degrees C for 2 minutes
 * 4) Place on ice for 2 minutes
 * 5) Add 700ul of LB
 * 6) Place at 37 degrees C for 40 minutes
 * 7) Spin down cells at 3,000-5,000 rpm
 * 8) Pipet off about 500ul
 * 9) Resuspend pellet in remainder.
 * 10) Plate ½ of the remainder (175ul)
 * 11) Save the other half in the fridge.
 * 12) If the plate doesn't have any colonies the following day, you can try plating the other half.