# Transforming chemically competent cells (Inoue)

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## Method

1. Thaw 25 - 200 μl TB buffer cells on ice. Do not use glass tubes, which adsorb DNA.
2. Add DNA, pipette gently to mix (keep volume of DNA less than 5% of the cell volume)
3. Incubate on ice for 30 minutes
• Note: If you are in a rush, you can shorten this incubation time to 5-10 min.
4. Incubate cells for 30 seconds at ${\displaystyle 42^{o}}$C.
5. Incubate cells on ice for 2 min.
6. Add 4 volumes of room temperature SOC (not critical)
7. Incubate for 1 hour at ${\displaystyle 37^{o}}$C on shaker.
• Note: Can also save some time here by reducing incubation to ~45 min.
• Note: Step can be eliminated if plating on Amp plates, but not most other antibiotics