Preparation of constructs with OmpA protein fusions

Michał K.

1. PCR on OmpA_linker and linker_alpha with OmpaL_N and AlphaP_XB primers (20 cycles, elongation length 1 min 15 s, annealing temperature 57°C).
2. PCR on OmpA_linker and linker_omega with OmpaL_N and OmegaP_EPB primers (20 cycles, elongation length 1 min 15 s, annealing temperature 57°C).
3. Gel electrophoresis of PCR products and gel-out of OmpA_alpha (1000 bp) and OmpA_omega (800 bp).
4. Electrophoresis to estimate the concentration of isolated DNA.
5. Overnight digest of purified OmpA_alpha and OmpA_omega DNA with NdeI and BamHI (Tango 2x buffer).
6. Inoculation of E. coli TOP10 with pACYC177 plasmid into liquid LB + kanamycin.

Blue/white and rifampicin test

Michał L., Ewa

Repetition of experiment from 11/06/2008.

Cloning of protein Z DNA to pET15b-OmpA-alpha in place of OmpA

Paweł

Ligation transformed into TOP10 strain and plated on LB + ampicillin.