Generator

Part:BBa_K942013:Design

Designed by: Luis Mario Leal Garza   Group: iGEM12_Tec-Monterrey   (2012-09-26)

pBAD+OmpA+RiAFP+Term


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 1205
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 1144
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 979
    Illegal AgeI site found at 1363
    Illegal AgeI site found at 1489
    Illegal AgeI site found at 1573
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI site found at 961


Design Notes

The RiAFP is better expressed when induced with 0.2-0.4% L-Arabinose. This part was designed by adding a 6xHis tag for affinity purification to the anti freeze protein from Raghium inquisitor and the signal sequence of the ompA gene for periplasmic secretion. This part was codon optimized for its expression in E. coli.

Source

The Raghium inquisitor Anti Freeze Protein (RiAFP) is a 137 AA protein that prevents the growth of ice crystals. This protein is produced natively in the beetle Raghium inquisitor. This part includes the OmpA signal sequence native from E.coli and the pBAD L-arabinose inducible promoter. (Kristiansen E, 2011)

References

Kristiansen, E. et all (2011) Structural characteristics of a novel antifreeze protein from the longhorn beetle Rhagium inquisitor. Insect Biochem Mol Biol. 41(2):109-17.