Week 12

From 2007.igem.org

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'''Testing our devices'''
'''Testing our devices'''
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We want to test if our newly-constructed devices work, that is if they beam flourescence when inducted with IPTG.
*In 2 different tubes we add 5ml of LB medium and 2 different colonies of [http://partsregistry.org/Part:BBa_I763019 I763019] plasmid.  
*In 2 different tubes we add 5ml of LB medium and 2 different colonies of [http://partsregistry.org/Part:BBa_I763019 I763019] plasmid.  
*In another tube we add 5ml of LB medium and a colony of PLac-cI-GFP 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.  
*After 2 hours we measure the OD value and it is around 0.5.  
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* For each original tube we divide the bacteria fluid in two different tubes.
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*For each original tube we divide the bacteria fluid in two different tubes.
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* We add the fluid without IPTG to one and the fluid with IPTG to the other one.
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*We add 1mM IPTG to the solution into one tube.
*The analyzed fluid without IPTG includes:
*The analyzed fluid without IPTG includes:
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-2.5ul of kanamicin,
-2.5ul of kanamicin,
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-50ul of IPTG.
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-50ul of 100mM IPTG.
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*We examine a bacteria fluid drop from of each tube with our fluorescence microscopy.  
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*We examine a bacteria fluid drop from of each tube with our fluorescence microscope.  
*The bacteria with [http://partsregistry.org/Part:BBa_I763019 I763019] plasmid don’t beam fluorescence with and without IPTG;  
*The bacteria with [http://partsregistry.org/Part:BBa_I763019 I763019] plasmid don’t beam fluorescence with and without IPTG;  
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*Very few bacteria with PLac-cI-GFP plasmid and without IPTG beam fluorescence;
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*Very few bacteria with PLac-cI-GFP plasmid with and without IPTG beam fluorescence.
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*Several bacteria with PLac-cI-GFP plasmid with IPTG beam fluorescence.  
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*After 3 hours we measured the OD value of each tubes and it is 1.4.
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In conclusion, we see that bacteria with [http://partsregistry.org/Part:BBa_I763019 I763019] plasmid don't seem to work at all, and bacteria with PLac-cI-GFP plasmid don't seem to work properly.
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*Bacteria are too concentrate, thus we dilute them.
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*We take 2ml of LB medium, 1ml of bacteria fluid with and without IPTG and 2ul of kanamicin.
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*Examining with fluorescence microscopy and we saw that only bacteria with PLac-cI-GFP plasmid beame fluorescence.
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Revision as of 09:06, 3 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 want to test if our newly-constructed devices work, that is if they beam flourescence when inducted with IPTG.

  • In 2 different tubes we add 5ml of LB medium and 2 different colonies of [http://partsregistry.org/Part:BBa_I763019 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 1mM IPTG to the solution into one tube.
  • 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 100mM IPTG.

  • We examine a bacteria fluid drop from of each tube with our fluorescence microscope.
  • The bacteria with [http://partsregistry.org/Part:BBa_I763019 I763019] plasmid don’t beam fluorescence with and without IPTG;
  • Very few bacteria with PLac-cI-GFP plasmid with and without IPTG beam fluorescence.

In conclusion, we see that bacteria with [http://partsregistry.org/Part:BBa_I763019 I763019] plasmid don't seem to work at all, and bacteria with PLac-cI-GFP plasmid don't seem to work properly.


09/21/07



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