Composite

Part:BBa_K2842669:Design

Designed by: Catherine Fan, Eloise Nee   Group: iGEM18_UCL   (2018-10-02)


mScarlet reporter with TerL-C intein on the N terminus


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NotI site found at 941
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 1224
    Illegal BsaI.rc site found at 28
    Illegal SapI.rc site found at 419
    Illegal SapI site found at 1130


Design Notes

Intein: The C terminal Acel-TerL intein was chosen to be used here as it’s N terminal regions is small (25 aa). As our construct can successfully be expressed even with the larger fusion it allows the smaller intein fragment to be used on the counterpart.

Reporter: The mScarlet protein was chosen due to it’s brightness and as it is known to be a fast folding protein making it an excellent reporter.

Modular design: SapI was chosen here as it leaves 3 bp overhangs simplifying the design of constructs as the overhang is maintained in frame.

Other design considerations:

Promotor: A T7 promotor (BBa_I712074) was chosen which can be induced with IPTG for strong expression.

Ribosome binding site: This RBS was generated using the sallis lab RBS calculator tool.

Regulatory elements: Contains a Lac operator which can be used to control leaky expression if needed.




Source

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References