Part:BBa_K3634005:Design

ATPG and NRPS Composite (E.coli Optimised)

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal NheI site found at 7
  Illegal NheI site found at 30
  Illegal NheI site found at 1561
  Illegal NheI site found at 1584
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 2059
  Illegal BamHI site found at 4112
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]

Design Notes

Due to the two-phase nature of our project, the selected promoter and RBS used may be subject to change after wet lab application in 2021 to maximise shinorine production. Golden gate assembly methods will facilitate this optimisation if necessary. It may also be required to include more than one single promoter for adequate expression of NRPS compared to the other enzymes involved in the shinorine pathway.

Source

The initial genomic sequence came from Anabaena variabilis ATCC 29413 which can be obtained from part K8140003 (Minnesota iGEM, 2012) before being optimised using the IDT optimisation tool. The promoter and RBS were selected from each respective Anderson catalogue based on their relative strength. The terminator BBa_B0015 sequence can also be taken from the iGEM registry.

References

Minnesota iGEM 2012 - https://parts.igem.org/Part:BBa_K814001

Minnesota iGEM 2012 - https://parts.igem.org/Part:BBa_K814003

Balskus, E.P, Walsh, C.T. 2010. The Genetic and Molecular Basis for Sunscreen Biosynthesis in Cyanobacteria. Science. 329 (5999): p1653-1656. DOI: 10.1126/science.1193637