Difference between revisions of "Part:BBa K2680051"
 
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===Mechanics===
 
===Mechanics===
  
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In this circuit, temperature-sensitive CI is expressed constitutively by J23107, a medium-weak constitutive promoter. This temperature-sensitive CI then represses the downstream CI promoter. Once the temperature is sufficiently raised, the ts-CI protein denatures, loosing its repression capabilities. Therefore, the CI promoter is freed, allowing for expression of the rapidly-maturing mScarlet-I. The mScarlet-I in this circuit has a pdt, a protein degradation tag, allowing for degradation by mf-Lon protease (not included in this circuit). Specifically, this circuit contains pdt#3, a strong degradation tag from the part series K2333401-K2333406 or K2333401.
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In this circuit, temperature-sensitive CI is expressed constitutively by J23107, a medium-weak constitutive promoter. This temperature-sensitive CI then represses the downstream CI promoter. Once the temperature is sufficiently raised, the ts-CI protein denatures, loosing its repression capabilities. Therefore, the CI promoter is freed, allowing for expression of the rapidly-maturing mScarlet-I. The mScarlet-I in this circuit has a pdt, a protein degradation tag, allowing for degradation by mf-Lon protease (not included in this circuit). Specifically, this circuit contains pdt#3, the strongest degradation tag from the part series K2333401-K2333406. To view a figure that displays the degradation rates of mScarlet-I with various pdt tags (relative to untagged mScarlet-I), see <partinfo>BBa_K2680251</partinfo>.
  
  
 
===Role in an IFFL System===
 
===Role in an IFFL System===
  
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This circuit provides the reporter (mscarlet-I-pdt#3) for an IFFL system. Specifically, this reporter is able to be degraded by mf-Lon. When these two circuits are combined into a single cells by having them both be on different plasmids, IFFL like behavior should be possible. However, through experimentation,  
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This circuit provides the reporter (mscarlet-I-pdt#3) for an IFFL system. Specifically, this reporter is able to be degraded by mf-Lon. When these two circuits are combined into a single cells by having them both be on different plasmids, IFFL like behavior should be possible. However, through experimentation, we have not identified an mf-Lon source strong enough to produce a pulse, that does not also kill the cells.
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<center>
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"https://static.igem.org/mediawiki/2018/e/ea/T--William_and_Mary--014_%2B_012_death.png"
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</center>
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For example, as can be seen in the above graph, when we try to combine a ts-CI mf-Lon on a psb3K3 plasmid with BBa_K2680051, the cells are killed upon dilution, and when the temperature is raised to 37C (grey region).
  
 
<!-- Add more about the biology of this part here
 
<!-- Add more about the biology of this part here

Latest revision as of 02:51, 18 October 2018


37C Temperature Sensitive mScarlet-I pdt#3

Temperature-Sensitive mScarlet-I pdt#3. Components: J23107 ts-CI + R0051 mScarlet-I pdt#3

Mechanics

In this circuit, temperature-sensitive CI is expressed constitutively by J23107, a medium-weak constitutive promoter. This temperature-sensitive CI then represses the downstream CI promoter. Once the temperature is sufficiently raised, the ts-CI protein denatures, loosing its repression capabilities. Therefore, the CI promoter is freed, allowing for expression of the rapidly-maturing mScarlet-I. The mScarlet-I in this circuit has a pdt, a protein degradation tag, allowing for degradation by mf-Lon protease (not included in this circuit). Specifically, this circuit contains pdt#3, the strongest degradation tag from the part series K2333401-K2333406. To view a figure that displays the degradation rates of mScarlet-I with various pdt tags (relative to untagged mScarlet-I), see BBa_K2680251.


Role in an IFFL System

This circuit provides the reporter (mscarlet-I-pdt#3) for an IFFL system. Specifically, this reporter is able to be degraded by mf-Lon. When these two circuits are combined into a single cells by having them both be on different plasmids, IFFL like behavior should be possible. However, through experimentation, we have not identified an mf-Lon source strong enough to produce a pulse, that does not also kill the cells.

"T--William_and_Mary--014_%2B_012_death.png"

For example, as can be seen in the above graph, when we try to combine a ts-CI mf-Lon on a psb3K3 plasmid with BBa_K2680051, the cells are killed upon dilution, and when the temperature is raised to 37C (grey region).

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
    Illegal NotI site found at 1518
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]