Week 12

From 2007.igem.org

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Line 1: Line 1:
::'''09/17/07'''
::'''09/17/07'''
*Ligations for:
*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_I763019 I763019];
-[http://partsregistry.org/Part:BBa_I763026 I763026] + [http://partsregistry.org/Part:BBa_I763004 I763004];
-[http://partsregistry.org/Part:BBa_I763026 I763026] + [http://partsregistry.org/Part:BBa_I763004 I763004];
Line 18: Line 18:
-[http://partsregistry.org/Part:BBa_I763028 I763028];
-[http://partsregistry.org/Part:BBa_I763028 I763028];
-
-Ptet-LacI-T-PLac-GFP;
+
-[http://partsregistry.org/Part:BBa_I763027 I763027];
-
-Ptet-LacI-GFP(Spe/Pst1), (Xba/Pst1).
+
-[http://partsregistry.org/Part:BBa_I763035 I763035] (Spe/Pst1), (Xba/Pst1).
*Digestion for:
*Digestion for:
-[http://partsregistry.org/Part:BBa_I763028 I763028] with Eco/Spe;
-[http://partsregistry.org/Part:BBa_I763028 I763028] with Eco/Spe;
-
-Ptet-LacI-T-PLac-GFP with Eco/Spe;
+
-[http://partsregistry.org/Part:BBa_I763027 I763027] with Eco/Spe;
-
-Ptet-LacI-GFP with Eco/Spe;
+
-[http://partsregistry.org/Part:BBa_I763035 I763035] with Eco/Spe;
-[http://partsregistry.org/Part:BBa_I763007 I763007] with Eco/Xba;
-[http://partsregistry.org/Part:BBa_I763007 I763007] with Eco/Xba;
-
-Plac-cI-LacY with Eco/Spe1;
+
-[http://partsregistry.org/Part:BBa_I763036 I763036] with Eco/Spe1;
*Band extraction from gel for all digestion and then we observe:
*Band extraction from gel for all digestion and then we observe:
-
-I763027, J04431 are died;
+
-[http://partsregistry.org/Part:BBa_I763028 I763028], [http://partsregistry.org/Part:BBa_I763027 I763027] are died;
-
-Ptet-LacI-GFP is correct;
+
-[http://partsregistry.org/Part:BBa_I763035 I763035] is correct;
-
-I763007 not found;
+
-[http://partsregistry.org/Part:BBa_I763007 I763007] not found;
-
-Plac-cI-LacY is correct.
+
-[http://partsregistry.org/Part:BBa_I763036 I763036] is correct.
::'''09/20/07'''
::'''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 [http://partsregistry.org/Part:BBa_I763031 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 [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 [http://partsregistry.org/Part:BBa_I7630231 I763031] 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 [http://partsregistry.org/Part:BBa_I763031 I763031] plasmid (tube C1, C2) don't seem to work properly either. We plan other experiments for next week.
Line 48: Line 70:
      
      
::'''09/21/07'''
::'''09/21/07'''
 +
Meeting: definition of fluorescence test protocol.
-
 
+
[[Bologna#Diary | Back]]
-
[[Bologna | Back]]
+

Latest revision as of 15:52, 26 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];

-[http://partsregistry.org/Part:BBa_I763027 I763027];

-[http://partsregistry.org/Part:BBa_I763035 I763035] (Spe/Pst1), (Xba/Pst1).

  • Digestion for:

-[http://partsregistry.org/Part:BBa_I763028 I763028] with Eco/Spe;

-[http://partsregistry.org/Part:BBa_I763027 I763027] with Eco/Spe;

-[http://partsregistry.org/Part:BBa_I763035 I763035] with Eco/Spe;

-[http://partsregistry.org/Part:BBa_I763007 I763007] with Eco/Xba;

-[http://partsregistry.org/Part:BBa_I763036 I763036] with Eco/Spe1;

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

-[http://partsregistry.org/Part:BBa_I763028 I763028], [http://partsregistry.org/Part:BBa_I763027 I763027] are died;

-[http://partsregistry.org/Part:BBa_I763035 I763035] is correct;

-[http://partsregistry.org/Part:BBa_I763007 I763007] not found;

-[http://partsregistry.org/Part:BBa_I763036 I763036] 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 [http://partsregistry.org/Part:BBa_I763031 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 [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 [http://partsregistry.org/Part:BBa_I7630231 I763031] 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 [http://partsregistry.org/Part:BBa_I763031 I763031] plasmid (tube C1, C2) don't seem to work properly either. We plan other experiments for next week.



09/21/07

Meeting: definition of fluorescence test protocol.


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