Difference between revisions of "Part:BBa I751501"

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sense TWO INPUTS, activation by AHL and repression by Lambda cI.
 
sense TWO INPUTS, activation by AHL and repression by Lambda cI.
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<span class='h3bb'>Sequence and Features</span>
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<partinfo>BBa_I751501 SequenceAndFeatures</partinfo>
  
 
===Usage and Biology===
 
===Usage and Biology===
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[[Image : Hybridgraph.JPG|thumb|300px| '''Fig.3: The result of assay0''' <br>Only in the presence of AHL and in the absence of lacI, the lux lac hybrid promoter is activated significantly.|right]]
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<br>
  
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<span class='h3bb'>Sequence and Features</span>
 
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<partinfo>BBa_I751501 SequenceAndFeatures</partinfo>
 
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<!-- Uncomment this to enable Functional Parameter display
 
 
===Functional Parameters===
 
===Functional Parameters===
 
<partinfo>BBa_I751501 parameters</partinfo>
 
<partinfo>BBa_I751501 parameters</partinfo>
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== [[Tokyo/AHL assay|1.AHL assay]] ==
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<br>[[Image:hill AHL.JPG|thumb|210px|'''Fig.1 The result of AHL assay''' Fluorescence intensity (arbitrary unit, a.u.) as a function of the concentration of AHL was determined. ]]
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'''Purpose'''
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<br>check how AHL activates lux-lac hybrid promoter.
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<br>These parameters(n2,K2) is decided.
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'''Result & Conclusion'''
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<br>As the concentration of AHL increases, GFP fluorescence increased, indicating that the hybrid promoter’s activation is strengthend with increasing concentration of AHL. From the activation graph in Fig. 1, the characteristics of the hybrid promoter expressed in Hill function, such as (n2,K2), is determined.
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<br>
 
<br>
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[[Image : Hybridgraph.JPG|thumb|300px| '''Fig.3: The result of assay0''' <br>Only in the presence of AHL and in the absence of lacI, the lux lac hybrid promoter is activated significantly.|right]]
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====⇒see more [http://parts.mit.edu/igem07/index.php/Tokyo/AHL_assay|details]====
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== [[Tokyo/IPTG assay|2.IPTG assay]] ==
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[[Image:hill IPTG.JPG|thumb|210px|'''Fig.2 The result of IPTG assay''' Fluorescence intensity as a function of the concentration of IPTG was determined. ]]
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'''Purpose'''
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<br>check how LacI represses this hybrid promoter.  In order to adjust LacI repression<!--IPTGはLacIの濃度を調節しているわけではないのでconcentrationではなくrepressionに変更-->, IPTG was added.
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<br>These parameters(n3,K3) is decided.
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<!--<br>To determine the order of the concentration of IPTG necessary for the activation of our lux-lac hybrid promoter in the LacI producing pTrc99A cells.
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<br> The order of the concentration is used for more detailed assay with narrower range of the IPTG concentration.-->
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'''Result & Conclusion'''
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<br><!--As the concentration of IPTG increases, GFP fluorescence increased, indicating that the hybrid promoter in the LacI expressing cell became increasingly strengthened by decreasing repression by LacI. The activation graph in Fig. 2, the characteristics of the hybrid promoter expressed in Hill function is determined.-->
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The concentration of IPTG dose-denpendently increased GFP fluorescence. It indicates that the hybrid promoter in the cells expressing LacI became increasingly strengthened by decline of repression by LacI. The activation graph in Fig. 2, the characteristics of the hybrid promoter expressed in Hill function is determined.  
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<br>
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====⇒see more [http://parts.mit.edu/igem07/index.php/Tokyo/IPTG_assay|details]====
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Revision as of 16:59, 26 October 2007

plux-cI hybrid promoter

sense TWO INPUTS, activation by AHL and repression by Lambda cI.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]

Usage and Biology

The characteristics of this hybrid promoter (Lux-lac hybrid promoter) have been determined though [http://parts.mit.edu/igem07/index.php/Tokyo/Works/Assay0| Assay 0]. The results are shown in Fig.3. The newly devised hybrid promoter is activated by AHL and repressed by LacI.
In the present project, this hybrid promoter has been tested on the BBa_I751100. (J54140del Pr)


Fig.3: The result of assay0
Only in the presence of AHL and in the absence of lacI, the lux lac hybrid promoter is activated significantly.


Functional Parameters

negative_regulators1
positive_regulators1

1.AHL assay


Fig.1 The result of AHL assay Fluorescence intensity (arbitrary unit, a.u.) as a function of the concentration of AHL was determined.

Purpose
check how AHL activates lux-lac hybrid promoter.
These parameters(n2,K2) is decided.

Result & Conclusion
As the concentration of AHL increases, GFP fluorescence increased, indicating that the hybrid promoter’s activation is strengthend with increasing concentration of AHL. From the activation graph in Fig. 1, the characteristics of the hybrid promoter expressed in Hill function, such as (n2,K2), is determined.


⇒see more [http://parts.mit.edu/igem07/index.php/Tokyo/AHL_assay|details]

2.IPTG assay

Fig.2 The result of IPTG assay Fluorescence intensity as a function of the concentration of IPTG was determined.

Purpose
check how LacI represses this hybrid promoter. In order to adjust LacI repression, IPTG was added.
These parameters(n3,K3) is decided.

Result & Conclusion
The concentration of IPTG dose-denpendently increased GFP fluorescence. It indicates that the hybrid promoter in the cells expressing LacI became increasingly strengthened by decline of repression by LacI. The activation graph in Fig. 2, the characteristics of the hybrid promoter expressed in Hill function is determined.

⇒see more [http://parts.mit.edu/igem07/index.php/Tokyo/IPTG_assay|details]