Composite

Part:BBa_K2957009:Design

Designed by: Miles Avery George   Group: iGEM19_MIT   (2019-10-19)


hEF1a, NeonGreen-C5a


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 3971
    Illegal XbaI site found at 4018
    Illegal SpeI site found at 3083
    Illegal PstI site found at 464
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 3971
    Illegal SpeI site found at 3083
    Illegal PstI site found at 464
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 3971
    Illegal BglII site found at 1932
    Illegal BamHI site found at 480
    Illegal XhoI site found at 2048
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 3971
    Illegal XbaI site found at 4018
    Illegal SpeI site found at 3083
    Illegal PstI site found at 464
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 3971
    Illegal XbaI site found at 4018
    Illegal SpeI site found at 3083
    Illegal PstI site found at 464
    Illegal AgeI site found at 81
    Illegal AgeI site found at 4024
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 3611
    Illegal SapI.rc site found at 3584


Design Notes

We used Modular Cloning Golden Gate Type II Assembly, so we had to consider how parts came together. We also used the natural secretion tag given in the part so we could use it to test chemokine secretion success by HEKs. We also had to consider which terminal the fluorescence/protein tag was added so as not to render the chemokine dysfunctional. See more on the MIT iGEM 2019 wiki: https://2019.igem.org/Team:MIT


Source

Parts of this part were ordered from IDT/Twist and then assembled with other parts (from iGEM/parts in our parts collection/from the Weiss lab) into this plasmid.

References