Difference between revisions of "Part:BBa K206000:Design"
 
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__NOTOC__
 
__NOTOC__
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<partinfo>BBa_K206000 short</partinfo>
 
  
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{|
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|<div style="align: center; valign: center; font-family: Arial; font-size: 12pt">
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{|
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|-
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|width="60px"|''Name'':
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|pBAD strong
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|-
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|width="60px"|''Input'':
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|width="100px"|[http://openwetware.org/wiki/Arabinose L-arabinose]
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|-
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|''Output'':
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| PoPS
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|}
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</div>
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<hr width="800px">
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<div class="noprint" style="padding: 10px; color: #ffffff; background-color: #C0C0C0; width: 800px; align: center">
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<center>
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[[Part:BBa_K206000 |Part Main Page]] &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
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[[Part:BBa_K206000:Design |Part Design]] &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
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[[Part:BBa_K206000:Characterization |Characterization]] &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
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[[pBAD Promoter Family |Family]] &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
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[[Part:BBa_K206000:Experience |Add Data]] &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
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</center>
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</div>
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<br>
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<br>
 
<partinfo>BBa_K206000 SequenceAndFeatures</partinfo>
 
<partinfo>BBa_K206000 SequenceAndFeatures</partinfo>
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<br>
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==Design Notes==
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Niland et al. found that certain mutations in the AraI1 operator site increased binding of the DNA to the AraC protein [[Part:BBa_K206000:Design#References|[1]]]. We applied all of these mutations with the goal of creating a stronger version of the pBAD promoter. See [[pBAD Promoter Family]] for more details.
  
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==Source==
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Site-directed mutagenesis was performed on <partinfo>I13453</partinfo> using the following primers:<br>
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Forward: 5'-PO4-TAATCTTATGGACTATCTTGCTATGGCATAGC-3'<br>
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Reverse: 5'-PO4-GCGGATCCTACCTGACGCTTTTTATC-3'
  
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===Design Notes===
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''Site-directed mutagenesis:'' We used the Finnzyme Phusion Site-directed Mutagenesis Kit in accordance with the manufacturer's directions.
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There were no special design considerations.
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<br>
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''Primers: ''Phosphorylated oligos were purchased from Integrated DNA Technologies (IDT) and resuspended in water.
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<br>
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''Template:'' <partinfo>I13453</partinfo> was obtained from the 2009 iGEM Spring Distribution according to Registry instructions and used as the template for site-directed mutagenesis.
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===Source===
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Site-directed mutagenesis was performed on <partinfo>I13453</partinfo> using the following primers:<br>
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Forward: TAATCTTATGGACTATCTTGCTATGGCATAGC<br>
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Reverse: GCGGATCCTACCTGACGCTTTTTATC
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===References===
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==References==
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Niland P, Hühne R, Müller-Hill B. (1996). How AraC Interacts Specifically with its Target DNAs. ''J. Mol. Biol.'' 264:667-674. [http://www.ncbi.nlm.nih.gov/pubmed/8980677]
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[http://www.ncbi.nlm.nih.gov/pubmed/8980677 [1]]  Niland P, Hühne R, and Müller-Hill B. (1996). How AraC Interacts Specifically with its Target DNAs. J. Mol. Biol. '''264''':667-674.

Latest revision as of 23:57, 21 October 2009


Name: pBAD strong
Input: [http://openwetware.org/wiki/Arabinose L-arabinose]
Output: PoPS

Part Main Page        Part Design        Characterization        Family        Add Data       




Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 125
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 65
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

Niland et al. found that certain mutations in the AraI1 operator site increased binding of the DNA to the AraC protein [1]. We applied all of these mutations with the goal of creating a stronger version of the pBAD promoter. See pBAD Promoter Family for more details.

Source

Site-directed mutagenesis was performed on BBa_I13453 using the following primers:
Forward: 5'-PO4-TAATCTTATGGACTATCTTGCTATGGCATAGC-3'
Reverse: 5'-PO4-GCGGATCCTACCTGACGCTTTTTATC-3'

Site-directed mutagenesis: We used the Finnzyme Phusion Site-directed Mutagenesis Kit in accordance with the manufacturer's directions.
Primers: Phosphorylated oligos were purchased from Integrated DNA Technologies (IDT) and resuspended in water.
Template: BBa_I13453 was obtained from the 2009 iGEM Spring Distribution according to Registry instructions and used as the template for site-directed mutagenesis.

References

[http://www.ncbi.nlm.nih.gov/pubmed/8980677 [1]] Niland P, Hühne R, and Müller-Hill B. (1996). How AraC Interacts Specifically with its Target DNAs. J. Mol. Biol. 264:667-674.