Team:Warsaw/Calendar-Main/25 June 2008

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Change of the reporter from pZC320 with B-galactosidase to GFP or RFP

Piotr, Weronika

  1. Inoculation of 10ml LB with E.coli TOP10 carrying pZC
  2. Isolation and chemotransformation with pSB1A2 standard plasmids carrying parts: BBa_E0840(GFP genetrator) and BBa_J04450 (RFP generator)
  3. Plating of chemotransformants on LB+ampicillin

Preparation of constructs: OmpA_alpha and OmpA_omega

Michał K.

Repetition of PCRs and gel-out purifications from 16 June.

Preparation of alpha-A and omega-A fusions

Michał L. Marcin and Ewa

  1. We've run gradient PCR to obtain Alpha-link and link-A.
    • DNA template: pDRIVE-TAPTAG
    • Taq polymerase
    • buffer suplemented with (NH4)2SO4
    • MgCl2 concentration gradient (from 1 to 4 mM)
    • annealing temp 55°C
    • elongation time 1 - 2 minutes
  2. We have successfully amplified Alpha-link and link-A (Fig. 1). Now it's time for PCL (Polymerase Chain Ligation) to create fusions Alpha-A and Omega-A.
Fig. 1. PCR product - link-A.