Transformation of E. coli

Materials required

• Appropriate E. coli strains (200 µl)
• Appropriate vector DNA
• Heat block
• Bucket of ice
• Pipettes
• Eppendorf tubes
• 1.5% agar plate containing appropriate antibiotics

Procedures

1. Thaw a 200 µl aliquot of the desired strain of E. coli and add the transforming DNA (10 ng of vector DNA in 10 µl).
2. Incubate on ice for 30 minutes.
3. Heat-shock the cells for 120 seconds at 42°C, and place on ice again for 3-4 minutes.
4. Add 1 ml of LB broth and incubate the cells at 37°C for 1-15 hr in a water bath with gentle shaking.
5. Plate 200 µl of transformed E. coli onto 1.5% agar plate containing the appropriate selection markers.
6. Incubate the plates overnight at 37°C.

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