Difference between revisions of "Part:BBa K2680057"

 
 
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<partinfo>BBa_K2680057 short</partinfo>
 
<partinfo>BBa_K2680057 short</partinfo>
  
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Tight Repression 37C Temperature-Sensitive mf-lon ssra.
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Components: J23100 ts-CI+ R0051 mf-Lon ssrA.
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===Mechanics===
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Strong promoter J23100 constitutively expresses the CI repressor protein, which binds the downstream CI promoter. However, this circuit contains temperature sensitive CI, which denatures at 37C and consequently loses its ability to bind the CI promoter. Therefore, increasing the temperature to 37C alleviates the repression of the CI promoter and allows for the expression of mf-Lon protease. An ssra degradation tag follows the mf-Lon in this circuit, increasing mf-Lon turnover.
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===Role in IFFL===
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In an IFFL (incoherent feed-forward loop), an input activates a reporter and its inhibitor. This circuit produces the mf-Lon protease, which can degrade and thus inhibit promoters like mScarlet-I. Thus, for use in an IFFL, this circuit must be combined with a reporter-containing circuit. A degradation tag, such as a pdt tag, should follow the reporter coding sequence.
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===Characterization===
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<html><img src="https://static.igem.org/mediawiki/parts/2/28/T--William_and_Mary--BBa_K2680057pulse.png"width=550px"/></html>
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As shown in the figure above, an IFFL composed of BBa_K2680057 and BBa_K333424 (pTet mscarlet-I-pdt#3) containing circuit created a pulse of fluorescence when chemically induced.
  
 
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Latest revision as of 01:46, 18 October 2018


Tight Repression 37C Temperature-Sensitive mf-lon ssra

Tight Repression 37C Temperature-Sensitive mf-lon ssra. Components: J23100 ts-CI+ R0051 mf-Lon ssrA.

Mechanics

Strong promoter J23100 constitutively expresses the CI repressor protein, which binds the downstream CI promoter. However, this circuit contains temperature sensitive CI, which denatures at 37C and consequently loses its ability to bind the CI promoter. Therefore, increasing the temperature to 37C alleviates the repression of the CI promoter and allows for the expression of mf-Lon protease. An ssra degradation tag follows the mf-Lon in this circuit, increasing mf-Lon turnover.

Role in IFFL

In an IFFL (incoherent feed-forward loop), an input activates a reporter and its inhibitor. This circuit produces the mf-Lon protease, which can degrade and thus inhibit promoters like mScarlet-I. Thus, for use in an IFFL, this circuit must be combined with a reporter-containing circuit. A degradation tag, such as a pdt tag, should follow the reporter coding sequence.

Characterization

As shown in the figure above, an IFFL composed of BBa_K2680057 and BBa_K333424 (pTet mscarlet-I-pdt#3) containing circuit created a pulse of fluorescence when chemically induced.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 51
    Illegal NheI site found at 74
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 2903
    Illegal AgeI site found at 2987
    Illegal AgeI site found at 3193
    Illegal AgeI site found at 3218
  • 1000
    COMPATIBLE WITH RFC[1000]