Signalling

Part:BBa_K2033001:Design

Designed by: Brady Dennison   Group: iGEM16_Arizona_State   (2016-08-15)
Revision as of 19:36, 13 September 2016 by Bpdennis (Talk | contribs)


C(12)-HSL Receiver Device - AubR


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal PstI site found at 395
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal PstI site found at 395
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal PstI site found at 395
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal PstI site found at 395
    Illegal AgeI site found at 205
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 184


Design Notes

Here is the inducible promoter sequence:

Stage 1-Insert Regulator ORFs

Phusion PCR

The AubR part was first synthesized by Ryan Muller in Spring of 2015. Stage 1 consisted of the insertion of the open reading frame into a cloning vector found here.

Caption
Reagent Volume(uL) Mix(x5)
DNA Plasmid 1.0 5.0
10uM F Primer 1.0 5.0
10uM R Primer 1.0 5.0
10mM dNTPs 1.0 5.0
Phusion Pol. 0.5 2.5
5x HF buffer 10 50
dH2O 36 180
Total 50.5

Stage 2-Insert Promoters

Source

This HSL receiver was recovered from a metagenomic project on soil. The specific source is unknown.

References

(1) Nasuno, E., N. Kimura, M. J. Fujita, C. H. Nakatsu, Y. Kamagata, and S. Hanada. "Phylogenetically Novel LuxI/LuxR-Type Quorum Sensing Systems Isolated Using a Metagenomic Approach." Applied and Environmental Microbiology 78.22 (2012): 8067-074. Web