Composite

Part:BBa_K2680050

Designed by: Stephanie Do, Xiangyi Fang, Ethan Jones, Jessica Laury, Adam Oliver, Lillian Parr, Chengwu Shen, Tinh Son, Julia Urban, Yashna Verma, Hanmi Zou   Group: iGEM18_William_and_Mary   (2018-10-06)


J23107 ts-CI+ R0051 mf-Lon

Design

This part is a j23107 ts-cI repressed mf-Lon. For more information on mf-Lon, and to see the source of the mf-Lon, see BBa_K2333011

Mechanics

In this circuit, temperature-sensitive CI is expressed 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 mf-Lon.

Characterization

Mf-Lon is capable of degrading proteins tagged with a specific pdt tag. If used in combination with such a tagged protein, an IFFL circuit can be produced. During our research, we utilized BBa_K2680050 as our initial heat sensitive mf-Lon. After experimentation, we found that the amount of mf-Lon produced was too high to create an IFFL circuit, as either not enough mscarlet-I could be produced to generate a pulse, or the amount of mf-Lon and mscarlet-I was too high, causing cell death (see the figure below).

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


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]


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Categories
Parameters
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