Difference between revisions of "Part:BBa K1132022"
 
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The couple TetR-pTet system was already described by  iGEM08_EPF-Lausanne. This construction is a first step to characterize the entire system. A constitutive promoter (BBa_J23116) was assemble to tetR and then assemble to pTet-rbs-RFP-term. pTet is a constitutive promoter. TetR is an inducer that binds to pTet promoter and thus, represses expression the downstream system. Supply of tetracycline or its analog aTc (anhydrotetracycline) is known to bind to tetR and invert the operation (inhibits expression of mRFP in this case).  
 
The couple TetR-pTet system was already described by  iGEM08_EPF-Lausanne. This construction is a first step to characterize the entire system. A constitutive promoter (BBa_J23116) was assemble to tetR and then assemble to pTet-rbs-RFP-term. pTet is a constitutive promoter. TetR is an inducer that binds to pTet promoter and thus, represses expression the downstream system. Supply of tetracycline or its analog aTc (anhydrotetracycline) is known to bind to tetR and invert the operation (inhibits expression of mRFP in this case).  
  
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<br><br>After 18 hours, clones containing the plasmid (BBa_J23116-TetR-pTet-RFP) show a leaky basal expression of mRFP. We suppose that the promoter was too weak to express TetR in large quantity. Assembly of a stronger promoter could improve the system and lock the response to an ON/OFF response.
 
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<br><br>Besides, an experience was done to analyze the effect of the aTc inducer. Result show a visible induction of the red fluorescent protein expression by addition of aTC (60 ng/mL).
 
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https://static.igem.org/mediawiki/2013/d/de/2022.png
 
  
 
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<partinfo>BBa_K1132022 parameters</partinfo>
 
<partinfo>BBa_K1132022 parameters</partinfo>
 
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===Results of BBa_K1132022 characterization===
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After 18 hours, clones containing the plasmid (BBa_J23116-TetR-pTet-RFP) show a leaky basal expression of mRFP. We suppose that the promoter was too weak to express TetR in large quantity. Assembly of a stronger promoter could improve the system and lock the response to an ON/OFF response.
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<br><br>Besides, an experience was done to analyze the effect of the aTc inducer. Result show a visible induction of the red fluorescent protein expression by addition of aTC (60 ng/mL).
 +
 +
https://static.igem.org/mediawiki/2013/d/de/2022.png

Latest revision as of 03:52, 5 October 2013

BBa_J23116-TetR-pTet-RFP

The couple TetR-pTet system was already described by iGEM08_EPF-Lausanne. This construction is a first step to characterize the entire system. A constitutive promoter (BBa_J23116) was assemble to tetR and then assemble to pTet-rbs-RFP-term. pTet is a constitutive promoter. TetR is an inducer that binds to pTet promoter and thus, represses expression the downstream system. Supply of tetracycline or its analog aTc (anhydrotetracycline) is known to bind to tetR and invert the operation (inhibits expression of mRFP in this case).


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 7
    Illegal NheI site found at 30
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 1526
    Illegal AgeI site found at 1638
  • 1000
    COMPATIBLE WITH RFC[1000]


Results of BBa_K1132022 characterization

After 18 hours, clones containing the plasmid (BBa_J23116-TetR-pTet-RFP) show a leaky basal expression of mRFP. We suppose that the promoter was too weak to express TetR in large quantity. Assembly of a stronger promoter could improve the system and lock the response to an ON/OFF response.

Besides, an experience was done to analyze the effect of the aTc inducer. Result show a visible induction of the red fluorescent protein expression by addition of aTC (60 ng/mL).

2022.png