Difference between revisions of "Part:BBa K2940010:Design"

 
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===Design Notes===
 
===Design Notes===
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This part is designed for the DyP-Cytb5 expression. So the signal peptides were removed from the original coding sequence to make sure DyP can anchor with Cytb5. The strong promoter and RBS were introduced to make the DNA construct. It was cloned into pSB1C3 using classcial cloning and was verified using ABTS assay
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This part is designed for the DyP-Cytb5 expression. So the signal peptides were removed from the original coding sequence (NC_000964.3) to make sure DyP can anchor with Cytb5. The strong constitute promoter and RBS combination (BBa_K608002) were inserted prior to each start codon. This part is related to the ghost shells project, which aims at immobilising DyP from Bacillus subtilis to the inner surface of the cytosolic membrane of Escherichia coli with the help of the C-terminal membrane anchor protein Cytb5. Then, the expression of Lysis protein E from PhiX174 can cause the formation of a pore in the cell wall of E. coli, which results in the release of the cytosol. Theoretically, the empty cellular envelopes with immoblised DyP can be used for degradation.
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===Source===
 
===Source===

Revision as of 14:55, 21 October 2019


Immobilised dye-degrading peroxidase: Dyp (without signal peptides)+ Cytochrome b5


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 7
    Illegal NheI site found at 30
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 995
    Illegal SapI site found at 425
    Illegal SapI site found at 783
    Illegal SapI.rc site found at 939


Design Notes

This part is designed for the DyP-Cytb5 expression. So the signal peptides were removed from the original coding sequence (NC_000964.3) to make sure DyP can anchor with Cytb5. The strong constitute promoter and RBS combination (BBa_K608002) were inserted prior to each start codon. This part is related to the ghost shells project, which aims at immobilising DyP from Bacillus subtilis to the inner surface of the cytosolic membrane of Escherichia coli with the help of the C-terminal membrane anchor protein Cytb5. Then, the expression of Lysis protein E from PhiX174 can cause the formation of a pore in the cell wall of E. coli, which results in the release of the cytosol. Theoretically, the empty cellular envelopes with immoblised DyP can be used for degradation.

Source

NC_000964.3


References