Reporter

Part:BBa_K2429035:Design

Designed by: Nia Myrie   Group: iGEM17_MIT   (2017-10-24)
Revision as of 23:39, 24 October 2017 by Nmyrie (Talk | contribs) (→‎Design Notes)

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phEF1a 3 Exon mKate-HBG Reporter


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 2944
    Illegal XbaI site found at 2349
    Illegal PstI site found at 315
    Illegal PstI site found at 820
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 2944
    Illegal PstI site found at 315
    Illegal PstI site found at 820
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 2944
    Illegal BglII site found at 569
    Illegal BamHI site found at 1183
    Illegal XhoI site found at 968
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 2944
    Illegal XbaI site found at 2349
    Illegal PstI site found at 315
    Illegal PstI site found at 820
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 2944
    Illegal XbaI site found at 2349
    Illegal PstI site found at 315
    Illegal PstI site found at 820
    Illegal NgoMIV site found at 703
    Illegal AgeI site found at 81
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI.rc site found at 1237


Design Notes

This part is human-codon optimized. We found a BsaI site in the middle of mKate Exon 2, so we use site-directed mutagenesis to change the base pair at position 1450 from a G to an A. We also mutated the HBG Intron 2 at position 888 from a C to an A because the paper we were referencing had mutated the HBG intron in the same way.

Source

The human beta globin introns came from the genome of HEK cells, and mKate from jellyfish.


References