Team:Warsaw/Calendar-Main/2 July 2008

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Preparation of constructs with OmpA protein fusions

  1. PCR on OmpA_linker and linker_alpha with OmpaL_N and AlphaP_XB primers (20 cycles, annealing 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, annealing length 1 min 15 s, annealing temperature 57°C).
  3. Gel electrophoresis of PCR products and gel-out of OmpA_alpha and OmpA_omega.
  4. Overnight digest of purified OmpA_alpha and OmpA_omega DNA with NdeI and BamHI.
  5. Overnight digest of pET15b plasmid with NdeI and BamHI, dephosphorylation with CIAP.