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Coding

Part:BBa_K1351036:Design

Designed by: Nikolai Peschek   Group: iGEM14_LMU-Munich   (2014-10-04)
Revision as of 19:30, 16 October 2014 by NikolaiP (Talk | contribs) (→‎Source)

Quorum sensing two component system (AIP-II sensing histidine kinase agrC, response regulator agrA)


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 1139
    Illegal AgeI site found at 1114
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 475


Design Notes

This part contains the native sequences of agrC-II and agrA from Staphylococcus aureus N315. Since these two parts are forming a two component system, this is actually a composite part. The basic parts agrCII and agrA can be yielded by using the NgoMIV or AgeI restriction sites with appropiate restriction sites in the pre- or suffix.

Source

This part is derived from Staphylococcus aureus N315 gDNA by PCR using primers with modified RFC25 prefix and suffix:

agrA-fwd: gatcgaattccgcggccgcttctagataaggaggagccggcATGAAAATTTTCATTTGCGAAGACG agrA-rev: gatcctgcagcggccgctactagtattaaccggtTATTTTTTTAACGTTTCTCACCG agrC-II-fwd: gatcgaattccgcggccgcttctagataaggaggagccggcTTGATTCCAACTTTTTCATCTATC agrC-II-rev: gatcctgcagcggccgctactagtattaaccggtGTTGTTAATAATTTCAACTTTTTGAATAAAG

The two intermediated parts agrA and agrC-II were then fused by using the restriction sites EcoRI and SpeI for agrA, as well as XbaI and PstI for agrC-II, and ligation into EcoRI and PstI cut pSB1C3, thus creating the composite part agrC-IIA.

References