Part:BBa_K3866031

Propionate Production Construct

Usage and Biology

This composite part consists of the following two Composite Parts:

sbm:stuffer: BBa_K3866029 & ygfG:ygfH: BBa_K3866030

Design Notes

The coding sequences were domesticated. We removed BsmBI and BsaI sites in order to be compatible with GoldenBraid and MoClo. The sequences are cloned in seva ω2 vector and have overhangs compatible for GoldenBraid cloning.

Figure 1. The ω module of the Propionate Production: α1:ParaBAD:RBS-sbm-Double terminator:ParaBAD:RBS-ygfG-Double terminator:ParaBAD:RBS-ygfH-Double terminator

Verification of Cloning

Figure 2.: (C=Cut, U=Uncut) Restriction digestion of α1-sbm-ygfG-ygfH (C5-C6-C7-C8-C9) with: EcoRV, Expected bands: 5727bp, 2167bp, 1509bp, 1434bp, 745bp, Positive result: C5-C6-C7-C8-C9

Figure 3.: (C=Cut, U=Uncut) Restriction digestion of α1-sbm-ygfG-ygfH with: BamHI, Expected bands: 5727bp, 2612bp, 1903bp, 1076bp, 264bp, Positive result: C1-C2-C3-C4-C5

Experimental Use and Experience

This part showed functionality at part BBa_K3866031

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 4655
  Illegal BamHI site found at 1148
  Illegal BamHI site found at 1424
  Illegal BamHI site found at 3343
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal AgeI site found at 983
  Illegal AgeI site found at 3178
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal SapI site found at 965
  Illegal SapI site found at 3160
  Illegal SapI.rc site found at 1883
  

References

Akawi L, Srirangan K, Liu X, Moo-Young M, Perry Chou C. Engineering Escherichia coli for high-level production of propionate. J Ind Microbiol Biotechnol. 2015 Jul;42(7):1057-72. https://doi.org/10.1007/s10295-015-1627-4