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Coding

Part:BBa_K1351036:Design

Designed by: Nikolai Peschek   Group: iGEM14_LMU-Munich   (2014-10-04)
Revision as of 19:29, 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, and ligation into EcoRI and PstI cut pSB1C3, thus creating the composite part agrC-IIA.

References