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;  
+
::-2.5ml 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 I763019 plasmid doesn’t beam fluorescence with (A2, B2) nor without (A1, B1)  IPTG;  
+
::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:

-[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

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 [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.
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 [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.
  • 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.



09/21/07



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