Week 3

From 2007.igem.org

  • We amplify some bio-bricks of interest to our project from the IGEM plates. We re-suspend and transform:
    • J04430 to test the GFP fluorescence detection with our experimental set-up;
    • J04431 to test the GFP (+LVA) half-life since we need a GFP with a short one for our project;
    • J04500, the PLac promoter inducible by IPTG;
    • J04631, the GFP (+LVA) protein.
  • We strake on plates with the right antibiotic.

  • We perform a test for GFP induction with IPTG. We pick a colony from the J04430 and J04431 plates and grow it in 5ml of LB medium during the day. In the afternoon we dilute the 5 ml cultures in 50ml and let them grow overnight.
  • We pick a colony from 07/17 plates, inoculate each one in 5ml of LB medium and incubate overnight.

  • In the morning we grow the J04430 and J04431 cultures to an OD = 0.4 – 0.6. We add 1mM IPTG to induce GFP expression to test fluorescence after 2h. (photos).
  • Although J04431 works correctly in presence of IPTG, J04430 doesn't. So, we perform a run on electrophoresis gel for the J04430 plasmid. We find that it doesn't match the right molecular weight. We think that maybe there's something wrong with the plasmid.

  • Plasmid digestion (link):we digest J04500 with Spe1 and Pst1 and J04631 with Xba1 and Pst1.
  • We perform the gel extraction procedure and store at -20°C.
  • We amplify I13507 and R0051from the IGEM plates.