- Ligated sequential and double digests of T9002 to 3K3
- Transforming in MG1655
- Re-doing BB PCR with vent and phusion
- Mix for rxn (100uL):
- 8uL 2.5mM dNTP
- 20uL phusion buffer
- 1.5uL f primer
- 1.5uL r primer
- 1uL DNA
- 67uL H2O
- Just realized that we skipped the ligation step completely (3K3 and 1AK3).
- Doing that now, discarding PCR product.
- Re-culturing from plate
- Checked the sequences; none of them have the promoter.
- Waiting for primers to insert the promoter.