Part:BBa_K2680066
39C Temperature-Sensitive (ptplA) mScarlet-I pdt#3b
Components: J23107 TPL + ptplA B0034 mScarlet-I pdt#3b
Mechanics
J23107, a medium-weak constitutive promoter, constitutively expresses TlpA39. Unlike the original TlpA, TlpA39 undergoes a conformational change at 39C and loses its ability to bind the downstream pTlpA. Therefore, TlpA39 halts its repression of promoter pTlpA at temperatures above 39C, inducing the expression of the rapidly-maturing mScarlet-I. The mScarlet-I in this circuit has a pdt degradation tag, allowing for degradation by the mf-Lon protease. Specifically, this circuit contains pdt#3b, a medium-strong degradation tag from 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.
As an IFFL
In this circuit, heat activates a reporter, mScarlet-I. In a complete IFFL (incoherent feed forward loop), an input must activate both a reporter and its inhibitor. Therefore, this circuit requires mf-Lon (or a different inhibitor of mScarlet-I) to act as an IFFL.
Characterization
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In this figure, heat induces both BBa_K268066 and a circuit containing mf-Lon, its inhibitor. The combined circuits thus act as an IFFL. However, the fluorescence in this figure continues to increase rather than making the pulse desired by Team William and Mary. William and Mary attributes this continued increase of fluorescence to the weakness of mf-Lon in this particular circuit.
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Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 51
Illegal NheI site found at 74
Illegal NotI site found at 613
Illegal NotI site found at 1920 - 21INCOMPATIBLE WITH RFC[21]Illegal XhoI site found at 898
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 1201
- 1000COMPATIBLE WITH RFC[1000]
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