Transformation of wt swMb pT7-7 (Myoglobin)
- Remove wt swMb pT7-7 from freezer, thaw, and let sit at room temp for about a half hour.
- Take a sterile eppendorf tube and place it on ice.
- Mix 50μL of NovaBlue Competent E.coli with 5μL of the wt swMb pT7-7 in the cold, sterile tube.
- Incubate this mixture for 30 minutes on ice.
- Transfer the tube to a heat block at 42°C for 90 seconds, and then transfer the tube back to ice for 5 minutes.
- Add 200μL of SOC media to the cells/PCR product.
- Shake the mixture at 250rpm at 37°C for one hour.
- Spread 100μL of the mixture on an LB plate with 100μg/mL ampicillin.
- The LB plate was made with 0.875g LB, 0.7g Agar, and 35mL of distilled water. The mixture was autoclaved on a liquid cycle. When the mixture cooled to about 60°C 35μL of 100mg/mL ampicillin was added to it and it was poured into a sterile petri dish.
- Incubate the plate overnight (inverted) at 37°C.
Update 2/28/12: 11 colonies grew overnight on the plate