Week 12
From 2007.igem.org
- 09/17/07
- Ligations for:
-[http://partsregistry.org/Part:BBa_I763026 I763026] + [http://partsregistry.org/Part:BBa_I763019 I763019];
-[http://partsregistry.org/Part:BBa_I763026 I763026] + [http://partsregistry.org/Part:BBa_I763004 I763004];
-[http://partsregistry.org/Part:BBa_I763025 I763025] + [http://partsregistry.org/Part:BBa_J04031 J04031].
- We transform ligations and strake them on plates.
- 09/18/07
- We inoculate a colony for yesterday ligations in 5ml of LB medium O/N.
- 09/19/07
- Miniprep for:
-[http://partsregistry.org/Part:BBa_I763028 I763028];
-Ptet-LacI-T-PLac-GFP;
-Ptet-LacI-GFP(Spe/Pst1), (Xba/Pst1).
- Digestion for:
-[http://partsregistry.org/Part:BBa_I763028 I763028] with Eco/Spe;
-Ptet-LacI-T-PLac-GFP with Eco/Spe;
-Ptet-LacI-GFP with Eco/Spe;
-[http://partsregistry.org/Part:BBa_I763007 I763007] with Eco/Xba;
-Plac-cI-LacY with Eco/Spe1;
- Band extraction from gel for all digestion and then we observe:
-[http://partsregistry.org/Part:BBa_I763028 I763028], Ptet-LacI-T-PLac-GFP are died;
-Ptet-LacI-GFP is correct;
-[http://partsregistry.org/Part:BBa_I763007 I763007] not found;
-Plac-CI-LacY is correct.
- 09/20/07
Testing our devices
- In 2 different tubes we add 5ml of LB medium and 2 different colonies of I763019 plasmid.
- In another tube we add 5ml of LB medium and a colony of PLac-cI-GFP plasmid. *After 2 hours we measure the OD value and it is around 0.5.
- For each original tube we divide the bacteria fluid in two different tubes.
- We add the fluid without IPTG to one and the fluid with IPTG to the other one.
- The analyzed fluid without IPTG includes:
-2,5ml of original tube fluid,
-2,5ml of LB medium,
-2,5ul of kanamicin.
- The analyzed fluid with IPTG includes:
-2,5ml of original tube fluid,
-2,5ml of LB medium,
-2,5ul of kanamicin,
-50ul of IPTG.
- We examine a bacteria fluid drop from of each tube with our fluorescence microscopy.
- The bacteria with I763019 plasmid don’t beam fluorescence with and without IPTG;
- very few bacteria with PLac-cI-GFP plasmid and without IPTG beam fluorescence;
- several bacteria with PLac-cI-GFP plasmid with IPTG beam fluorescence.
- After 3 hours we measured the OD value of each tubes and it is 1,4.
- Bacteria are too concentrate, thus we dilute them.
- We take 2ml of LB medium, 1ml of bacteria fluid with and without IPTG and 2ul of kanamicin.
- Examining with fluorescence microscopy and we saw that only bacteria with PLac-cI-GFP plasmid beame fluorescence.
- 09/21/07
