Coding

Part:BBa_K4764001:Design

Designed by: Katie Spivakovsky   Group: iGEM23_MIT   (2023-10-08)


3xFLAG-IL6-mCherry fusion protein


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal XbaI site found at 549
    Illegal PstI site found at 529
    Illegal PstI site found at 604
    Illegal PstI site found at 625
    Illegal PstI site found at 673
    Illegal PstI site found at 1078
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal PstI site found at 529
    Illegal PstI site found at 604
    Illegal PstI site found at 625
    Illegal PstI site found at 673
    Illegal PstI site found at 1078
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal XbaI site found at 549
    Illegal PstI site found at 529
    Illegal PstI site found at 604
    Illegal PstI site found at 625
    Illegal PstI site found at 673
    Illegal PstI site found at 1078
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal XbaI site found at 549
    Illegal PstI site found at 529
    Illegal PstI site found at 604
    Illegal PstI site found at 625
    Illegal PstI site found at 673
    Illegal PstI site found at 1078
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI site found at 691


Design Notes

To choose how to order the components of the fusion protein, we had to consider the fact that 3xFLAG tag purification involves cleavage of the 3xFLAG portion off of the remainder of the protein; this forced us to put the tag on the 3' end. We also had to include short glycine linkers in between the 3 components in order to space out the components so that each protein could correctly fold independently of the others; this allowed IL-6 to retain its epitopes and mCherry to retain its red fluorescence.


Source

The sequence for IL-6 is publicly available. The sequence of the 3xFLAG tag is a short stretch of nucleotides, also publicly available. The mCherry sequence is publicly available as well.

References