Week 12

From 2007.igem.org

  • Ligations for:

-I763026 + I763019;

-I763026 + I763004;

-I763025 + J04031.

  • We transform ligations and strake them on plates.

  • We inoculate a colony for yesterday ligations in 5ml of LB medium O/N.

  • Miniprep for:



-I763035 (Spe/Pst1), (Xba/Pst1).

  • Digestion for:

-I763028 with Eco/Spe;

-I763027 with Eco/Spe;

-I763035 with Eco/Spe;

-I763007 with Eco/Xba;

-I763036 with Eco/Spe1;

  • Band extraction from gel for all digestion and then we observe:

-I763028, I763027 are died;

-I763035 is correct;

-I763007 not found;

-I763036 is correct.


Testing our devices

We wanted to test if our newly-constructed devices work, that is if they beam fluorescence when inducted with IPTG. To do so we performed 3 tests in parallel in 3 different tubes, that we name here A, B and C.

1. In tube A e in tube B we add 5ml of LB medium and 2 different colonies of I763019 plasmid.
2. In tube C we add 5ml of LB medium and a colony of I763031 plasmid.
3. After 2 hours we measure the OD value of tubes A, B, C and it is around 0.5.
4. For tubes A, B, C we divided the bacteria fluid in two different tubes A1, A2; B1, B2; C1, C2.
5. We add 1mM IPTG to the solution into tubes A2, B2, C2.
  • The analyzed fluid without IPTG (tubes A1, B1, C1) includes:
-2.5ml of original tube fluid;
-2.5ml of LB medium;
-2.5ul of kanamicin.
  • The analyzed fluid with IPTG (tube A2, B2, C2) includes:
-2.5ml of original tube fluid;
-2.5ml of LB medium;
-2.5ul of kanamicin;
-50ul of 100mM IPTG.
6. We examine a bacteria fluid drop of tubes A1, A2, B1, B2, C1, C2 with our fluorescence microscope.
7. The bacteria with I763019 plasmid doesn’t beam fluorescence with (A2, B2) nor without (A1, B1) IPTG;
8. Very few bacteria with I763031 plasmid with (C2) and without (C1) IPTG beam fluorescence.
  • In conclusion, we see that bacteria with I763019 plasmid (tubes A1, B1, A2, B2) don't seem to work at all, and bacteria with I763031 plasmid (tube C1, C2) don't seem to work properly either. We plan other experiments for next week.


Meeting: definition of fluorescence test protocol.