Intermediate

Part:BBa_K3128015:Design

Designed by: Melodie Dupre   Group: iGEM19_Grenoble-Alpes   (2019-09-18)
Revision as of 18:07, 1 October 2019 by Melo (Talk | contribs) (Source)


COMP fused with T18 subpart of Bordetella Pertussis adenylate cyclase under lactose promoter


Euromedex BACTH kit contains the pUT18 plasmid with a MCS cassette followed by the T18 subpart of the Bordetella pertussis adenylate cyclase. It allows the cloning of the protein of interest in fusion with the T18 subpart. To respect RFC rules, the plasmid has been amplificated by PCR using primers that bind from either site of the MCS cassette to eliminate it. Those primers contain also restriction sites enabling the cloning of the protein in the system. The plasmid was digested by the corresponding enzymes and the protein gene was inserted.

BBa K3128015 design1.png
BBa K3128015 design2.png


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 1465
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 1006
    Illegal NgoMIV site found at 1416
    Illegal AgeI site found at 1222
  • 1000
    COMPATIBLE WITH RFC[1000]

Source

pUT18 plasmid from Euromedex BACTH kit was used.
OmpX gene and GGS linker were synthesized by IDT because they were unavailable on iGEM plates.

References