Composite

Part:BBa_K4179008

Designed by: Yasmin Habib   Group: iGEM22_Technion-Israel   (2022-10-05)


PsPT1 under Rhlr promoter + P2A + mCherry reporter

This composite part comprises of an umbelliferone-6 prenyl transferase PsPT1 (BBa_K4179001), under the rhlr-regulated promoter (BBa_R0071). Downstream to PsPT1 there is a P2A sequence (BBa_K4179005), which is a self-cleavage element that induces ribosomal skipping during translation of a protein inside the cell. Downstream to which there is an mCherry (BBa_K106005) reporter gene, and an rrnB terminator sequence (BBa_K4047025).


Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 271
    Illegal EcoRI site found at 706
    Illegal XbaI site found at 2149
    Illegal PstI site found at 199
    Illegal PstI site found at 1789
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 271
    Illegal EcoRI site found at 706
    Illegal NheI site found at 1354
    Illegal PstI site found at 199
    Illegal PstI site found at 1789
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 271
    Illegal EcoRI site found at 706
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 271
    Illegal EcoRI site found at 706
    Illegal XbaI site found at 2149
    Illegal PstI site found at 199
    Illegal PstI site found at 1789
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 271
    Illegal EcoRI site found at 706
    Illegal XbaI site found at 2149
    Illegal PstI site found at 199
    Illegal PstI site found at 1789
    Illegal AgeI site found at 1428
  • 1000
    COMPATIBLE WITH RFC[1000]


Usage and Biology

In this genetic system, the mCherry expression is tied directly to the start codon of PsPT1, while not being covalently bound to it. In such a design the mCherry signal corresponds to a 1:1 ratio, at the very least, which allows for a lower estimate of PsPT1's expression.

PsPT1 enzyme is the first enzyme in the decursin biosynthesis pathway, which the team of Technion 2022 was attempting to clone into a bacterial system. For more information about the PsPT1 enzyme, visit its page in the registry (BBa_K4179001), or visit the team’s wiki page.


The team used this sequence in the rhlr-tdpp7-m/cherry plasmid, which already holds the rhlr gene.

[edit]
Categories
Parameters
None