Composite

Part:BBa_K4886006:Design

Designed by: Leyu Xu   Group: iGEM23_Nanjing-BioX   (2023-09-17)
Revision as of 15:03, 10 October 2023 by Xuleyu (Talk | contribs) (→‎Source)


Ptkt-P/Xpk(BD)


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 1675
    Illegal XbaI site found at 618
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 1675
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 1675
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 1675
    Illegal XbaI site found at 618
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 1675
    Illegal XbaI site found at 618
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

Our project aims to reduce the carbon emission during the fermentation of Clostridium tyrobutyricum (C. tyrobutyricum) by constructing NOG pathway to integrate with the native EMP pathway of the strain. NOG pathway is an artificial pathway and functions as a fructose-6-phosphate (F6P) shunt. Through carbon rearrangement, this pathway can convert 1mol of F6P to 3mol of AcP without any loss of carbon. By comparison, we found that most of the key enzymes in NOG pathway natively exist in C. tyrobutyricum and the functions of the absent key enzymes can be carried out by phosphoketolases (Figure 1). Therefore, in order to construct NOG pathway in C. tyrobutyricum L319, we introduced phosphoketolase (F/Xpk) gene into the strain. Ptkt promoter is a native transcriptional promoter in C. tyrobutyricum L319. We used Ptkt, Pthl and Pfba promoters to test which can provide the most robust expression of F/Xpk in the strain.

Figure 1 NOG pathway and related enzyme genes (red arrows indicate lack of key enzymes in Clostridium tyrobutyricum)

Source

The Ptkt promoter sequence is from BBa_K4886005.

The F/Xpk sequence is from BBa_K4119076.

The terminator sequence is from BBa_K3585002.

References