Deleted
Experience: None
Not Used
Get This Part
Plasmid

Part:BBa_M12068:Design

Designed by: Grant Robinson   Group: MIT BE 20.20 - S09 MIT BE 20.20 - S10   (2010-05-11)

BioBrick-Compatible Broad-Host-Range Vector pPMQAK1


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal prefix found in sequence at 1924
    Illegal suffix found in sequence at 2621
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 1924
    Illegal SpeI site found at 2622
    Illegal PstI site found at 2636
    Illegal NotI site found at 1930
    Illegal NotI site found at 2629
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 1924
    Illegal BamHI site found at 2609
    Illegal XhoI site found at 817
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal prefix found in sequence at 1924
    Illegal suffix found in sequence at 2622
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal prefix found in sequence at 1924
    Illegal XbaI site found at 1939
    Illegal SpeI site found at 2622
    Illegal PstI site found at 2636
    Illegal AgeI site found at 3681
    Illegal AgeI site found at 4082
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 963
    Illegal BsaI.rc site found at 2059


Design Notes

According to the authors' article, although BioBrick expression in non-Escherichia coli organisms was indeed possible with the use of the vector, there are issues with promoter expression (in particular, Plac and Ptet are essentially dysfunctional) and apparent issues with RNA Polymerase interactions with at least some protein-mediated activation/inhibition reactions; in particular, one designed promoter, Ptrc2O was properly repressed by LacI but was unable to return to constitutive activity with the addition of surplus IPTG. In short, protein expression via the pPMQAK1 shuttle vector is apparently viable, but dedicated promoters may well have to be designed depending on the chassis.


Source

http://www.ncbi.nlm.nih.gov/pubmed/20236988 [Location of the original article containing reference to pPMQAK1.]

http://www.ncbi.nlm.nih.gov/nuccore/292601742 [Location of the pPMQAK1 vector expressed here in GenBank.]

References