Part:BBa_K3773520:Design

Circuit to report flup2 expression in vivo

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 AgeI site found at 287
  
• 1000
  COMPATIBLE WITH RFC[1000]

Design Notes

Our analysis revealed that the flu gene is usually significantly downregulated when a circuit is introduced to an E. coli cell. This consistent downregulation means that we can use flu gene transcription levels to assess the effect of host-circuit interactions on the host cell. The flu gene controls biofilm production, specifically that of AG43¹, and we chose to use this gene as an indicator of cellular secretion being affected by introduction of a second, heterologous circuit.

The flu gene controlled by multiple promoters: flup, flup1, and flup2. flup activates both isrC and <i>flu, whereas flup1 and flup2 each only activate the flu gene²; therefore we eliminated the possibility of using flup as our promoter due to its lack of specificity (RegulonDB, 2019). We designed two versions of this circuit, with this version using flup2 as the promoter, and the other, K3773519, using flup1.

We place sfGFP after this promoter, allowing it to report expression.

The sequence inputted as a scar after the RBS is a spacer which has been shown to allow for expression with BBa_B0034 and sfGFP³.

UNS 1 and UNS 10 flank this part in order to allow for easy Gibson assembly as detailed by Torella et al., 2014⁴.

Source

See basic parts.