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I am making competent cells using DB3.1 E. coli cells. Even after following the exact protocol (Inoue method for ultracompetent cells) given in 'Sambrook and Russel', I am not getting transformation efficiency of more than 104. I am using a 5.1kb plasmid for checking transformation efficiency.

I will be thankful if any of you can share your experience in this experiment.

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How are you storing the cells? Competent cells can be very finicky. In my lab, we usually flash-freeze them with ethanol/dry-ice and store at -81 degrees (long-term use) or we store them surrounded by ice at 4 degrees (short-term use).

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