Maloof Lab:Cell Transformation

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E. coli (DH5α chemically-competent) Transformation Thaw one aliquot of DH5α competent cells (from –80°C freezer) on ice Add 5 μl DNA and 100 μl competent cells to new cell culture tube Incubate on ice for 30 minutes Heat shock in 42°C water bath for 30 seconds Cool on ice Add 900 μl liquid media (e.g. LB) to cell culture tube Shake at 37°C for 40-60 minutes Centrifuge at 5000 rpm for two minutes Remove all but 100-150 μl of the LB and resuspend pellet Spread 100-150 μl on plate of media + antibiotic (e.g. LB agar + carb) Incubate plates up-side-down, overnight at 37°C