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Team:Brown/Notebook/June29
From 2010.igem.org
Tuesday, June 29 2010
Prepare to do competence protocol
Started overnight culture on 6/28 at around 6:30 PM.
Took 1 mL of cells, diluted in ~250 mL LB. Set in shaking incubator at 10:36 AM.
- By accident, we resuspended in the wrong amount of TfBI.
Put cells at -80°C
Did a test transformation to check if cells were actually competent or not.
Nanodrop reading of DNA: 11ng/µl Use 2 µl DNA for transformation for ~20ng for the transformation
Put plates in 37°C at 5:30 PM.
Followed Gary’s protocol:
- 100 µl cell suspension in pre-cooled centrifuge tube
- Add DNA ligation to cells, mix gently with pipette (do this all on ice).
- Incubate 20 minutes on ice.
- Heat shock at 42°C for 2 minutes.
- Back on ice for 2 minutes.
- Add 0.5 ml LB media
- Spread on amp+ plates
- 37°C overnight.
