Introducing plasmid DNA into bacteria

material: chemical competent cells, heatblock (42°C), shaker (37°) LB plates (with antibiotics), LB-Medium (without entibiotics), ice
duration: 2 h

1. add 1 µL plasmid DNA to 25 µL chemical competent cells (thaw on ice and handle gently)
2. incubate 10 min on ice
3. heat shock 45 sec 42°C
4. incubate 2 min on ice
5. add 500 µL LB medium (without antibiotics)
6. incubate 1 h, 37°C (for re-trafo 15 min is sufficient)
7. centrifuge 4000 rpm, 1min (discard supernatant, resuspend in remaining medium)
8. plate 15 µL on LB-plates with antibiotics
9. incubate over night at 37°C

1. add 5 µL Gibson mix to 25 µL chemical competent cells (thaw on ice and handle gently)
2. heat shock 45 sec 42°C
3. incubate 2 min on ice
4. add 500 µL LB medium (without antibiotics)
5. incubate 1 h, 37°C
6. plate 300 µL on LB-plates with antibiotics
7. incubate over night at 37°C