Competition tests

Piotr

1. Plasmid isolation from previous day's cultures.
2. Digest with SacI and BamHI.
3. Electrophoresis.

Conclusion: cells with interracting protein survive competition!

MutD5 testing

Emilia

Inoculation of MutD5 into medium with tetracycline.

Optimisation of primers for preparation of parts

Michał K.

1. PCR on pACYC177+OmpA_omega_deltaA_alpha plasmid using AL_BNXNE and APSacSpe primers (elongation length 45s) to obtain deltaA fragment.
2. PCR on pACYC177+OmpA_omega_deltaA_alpha plasmid using LinLSXNE and AlfaPSpe primers (elongation length 60s) to obtain link_alpha fragment.
3. PCR on pACYC177 + OmpA_omega plasmid using LinLSXNE and OmegPSpe primers (elongation length 60s) to obtain link_omega fragment.
4. PCR on pACYC177+OmpA_omega_deltaA_alpha plasmid using OmpLNXNE and LinP_BS primers (elongation length 90s) to obtain Omp_omega fragment.

Each reaction was carried out with 25 cycles and in temperature gradient from 55 to 75 °C (four reactions).

5. Gel electrophoresis of PCR products.
6. Repetition of PCR on pACYC177+OmpA_omega_deltaA_alpha plasmid using LinLSXNE and AlfaPSpe primers (elongation length 60s) to obtain link_alpha fragment. Reaction was carried out with 25 cycles and in temperature gradient from 55 to 75 °C (four reactions).
7. Gel electrophoresis of PCR products.