Composite

Part:BBa_K4387978:Design

Designed by: Nathalie Weibel   Group: iGEM22_UZurich   (2022-09-30)


Nitric Oxide Sensing Genetic Circuit


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 708
    Illegal BglII site found at 1287
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

An important part of our engineering cycle was the characterization of GFP expression when changing the amount of ribosome binding sites after the NO sensor. We found that the best solution with the highest GFP yields contains two different RBSs inserted right after the promoter pNorVß. For further information about the promoter, please go to the part BBa_K4387000.


Source

The sequence of the nitric oxide sensor was take from the paper by Xiaoyu J. Chen et al. [1] Nanobody sequence was obtained from the patent mentioned in the respective nanobody part registry (BBa_K4387996). [2] The source for the HlyA secretion system (BBa_K4387987) is from a paper by Ruano-Gallego et al. [3]


References

  • [1] Xiaoyu J. Chen et al., 2021, Rational Design and Characterization of Nitric Oxide Biosensors in E. coli Nissle 1917 and Mini SimCells
  • [2] Silence, Karen, Lauwereys, Marc, De Haard, Hans, et al. "Single domain antibodies directed against tumour necrosis factor-alpha and uses therefor", Int. Publication Number: WO 2004/041862 A2, 21 May 2004
  • [3] Ruano-Gallego, D., Fraile, S., Gutierrez, C. et al. Screening and purification of nanobodies from E. coli culture supernatants using the hemolysin secretion system. Microb Cell Fact 18, 47 (2019). https://doi.org/10.1186/s12934-019-1094-0