Repeat steps 14-16 for all remaining electroporations. Make sure you add the additional controls:
- No DNA negative control = for an idea of selection strength
- Uninduced (no arabinose) control with DNA = for an idea of background plasmid contamination electroporation. Especially important if you PCR cleaned-up your fragment.
- The following primer pair can be used as a positive control:
F GAAGCAGTTAAGCTAGGCGGATTGAAGATTCGCAGGAGAGCGAGCGAAAACTCACGTTAAGGGATTTTG
R ATCAGCCGGGTGGCAACTCTGCCATCCGGCATTTCCCCGCAAATGGCACTTTTCGGGGAAATGTG
These primers delete the UNG gene and insert the Kanamycin cassette. They do not have FRT sites and amplify well from any pET28 vector.