Week 12
From 2007.igem.org
(Difference between revisions)
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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. | 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. | + | ::1. In tube A e in tube B we add 5ml of LB medium and 2 different colonies of [http://partsregistry.org/Part:BBa_I763019 I763019] plasmid. |
::2. In tube C we add 5ml of LB medium and a colony of PLac-cI-GFP plasmid. | ::2. In tube C we add 5ml of LB medium and a colony of PLac-cI-GFP plasmid. | ||
::3. After 2 hours we measure the OD value of tubes A, B, C and it is around 0.5. | ::3. After 2 hours we measure the OD value of tubes A, B, C and it is around 0.5. | ||
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* The analyzed fluid without IPTG (tubes A1, B1, C1) includes: | * The analyzed fluid without IPTG (tubes A1, B1, C1) includes: | ||
::-2.5ml of original tube fluid; | ::-2.5ml of original tube fluid; | ||
| - | :: | + | ::-2.5ml of LB medium; |
::-2.5ul of kanamicin. | ::-2.5ul of kanamicin. | ||
*The analyzed fluid with IPTG (tube A2, B2, C2) includes: | *The analyzed fluid with IPTG (tube A2, B2, C2) includes: | ||
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::-2.5ml of LB medium; | ::-2.5ml of LB medium; | ||
::-2.5ul of kanamicin; | ::-2.5ul of kanamicin; | ||
| - | -50ul of 100mM IPTG. | + | ::-50ul of 100mM IPTG. |
::6. We examine a bacteria fluid drop of tubes A1, A2, B1, B2, C1, C2 with our fluorescence microscope. | ::6. We examine a bacteria fluid drop of tubes A1, A2, B1, B2, C1, C2 with our fluorescence microscope. | ||
| - | ::7. The bacteria with | + | ::7. The bacteria with [http://partsregistry.org/Part:BBa_I763019 I763019] plasmid doesn’t beam fluorescence with (A2, B2) nor without (A1, B1) IPTG; |
::8. Very few bacteria with PLac-cI-GFP plasmid with (C2) and without (C1) IPTG beam fluorescence. | ::8. Very few bacteria with PLac-cI-GFP 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 PLac-cI-GFP plasmid (tube C1, C2) don't seem to work properly either. We plan other experiments for next week. | + | *In conclusion, we see that bacteria with [http://partsregistry.org/Part:BBa_I763019 I763019] plasmid (tubes A1, B1, A2, B2) don't seem to work at all, and bacteria with PLac-cI-GFP plasmid (tube C1, C2) don't seem to work properly either. We plan other experiments for next week. |
Revision as of 15:13, 4 October 2007
- 09/17/07
- Ligations for:
- 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:
-I763028;
-Ptet-LacI-T-PLac-GFP;
-Ptet-LacI-GFP(Spe/Pst1), (Xba/Pst1).
- Digestion for:
-I763028 with Eco/Spe;
-Ptet-LacI-T-PLac-GFP with Eco/Spe;
-Ptet-LacI-GFP with Eco/Spe;
-I763007 with Eco/Xba;
-Plac-cI-LacY with Eco/Spe1;
- Band extraction from gel for all digestion and then we observe:
-I763028, Ptet-LacI-T-PLac-GFP are died;
-Ptet-LacI-GFP is correct;
-I763007 not found;
-Plac-CI-LacY is correct.
- 09/20/07
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 PLac-cI-GFP 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 PLac-cI-GFP 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 PLac-cI-GFP plasmid (tube C1, C2) don't seem to work properly either. We plan other experiments for next week.
- 09/21/07
