Composite

Part:BBa_K648019

Designed by: Jim Rose   Group: iGEM11_Penn_State   (2011-07-05)


Fast-Fusion GFP-LacZa Reporter (TEV cleavage with long linker)

This is a fast-acting reporter using the LacZa gene (BBa_K648012) fused to GFP (BBa_K648013). In its normal un-cleaved state the polymerization ability necessary for enzymatic action is inactivated. Once the fusion protein is cleaved by the TEV protease the B-gal enzyme encoded by the LacZa gene is able to convert X-gal to galactose and 5-bromo-4-chloro-3-hydroxyindole, the latter of which is oxidized to the dark blue product 5,5'-dibromo-4,4'-dichloro-indigo. This reporter is able to produce a visible blue color response in a matter of minutes.

This version uses the TEV cleavage site (BBa_K648008) followed by the smallest of the three flexible fusion protein linkers (BBa_K648005). The linker is attached to the N-terminus of the LacZa gene.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 7
    Illegal NheI site found at 30
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 704
    Illegal BsaI.rc site found at 871


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