Difference between revisions of "Part:BBa K4727000:Design"

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===Design Notes===
 
===Design Notes===
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The plasmid backbone has been deleted of five prohibited restriction sites, the region was substituted with the standard RFC[10] prefix and suffix. Between them has been placed the part <partinfo>BBa_J23119</partinfo>, a promoter that has been demonstated to be working in <i> A. baumannii </i> and <i> K. pneumoniae </i> [1]
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The plasmid backbone originally designed by Wang et al. has been deleted of five prohibited restriction sites, the region was substituted with the standard RFC[10] prefix and suffix [1].
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+
  
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Therefore, to establish a standardized backbone suitable for various experiments in both of these bacteria, we opted to proceed in stages.
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Since all the necessary restriction sites that needed to be eliminated were within a multiple cloning site, we decided to excise that portion and insert a newly synthesized fragment containing both the prefix and the suffix.
  
 
===Source===
 
===Source===

Latest revision as of 20:52, 11 October 2023


A. baumannii & K. pneumoniae standard backbone


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal prefix found at 3664
    Illegal suffix found at 3721
  • 12
    INCOMPATIBLE WITH RFC[12]
    Plasmid lacks a prefix.
    Plasmid lacks a suffix.
    Illegal EcoRI site found at 3664
    Illegal NheI site found at 969
    Illegal NheI site found at 3692
    Illegal NheI site found at 3715
    Illegal SpeI site found at 3722
    Illegal PstI site found at 3736
    Illegal NotI site found at 3670
    Illegal NotI site found at 3729
  • 21
    INCOMPATIBLE WITH RFC[21]
    Plasmid lacks a prefix.
    Plasmid lacks a suffix.
    Illegal EcoRI site found at 3664
    Illegal BglII site found at 1162
    Illegal XhoI site found at 1775
    Illegal XhoI site found at 3780
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal prefix found at 3664
    Illegal suffix found at 3722
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal prefix found at 3664
    Plasmid lacks a suffix.
    Illegal XbaI site found at 3679
    Illegal SpeI site found at 3722
    Illegal PstI site found at 3736
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Plasmid lacks a prefix.
    Plasmid lacks a suffix.
    Illegal SapI site found at 4008


Design Notes

The plasmid backbone originally designed by Wang et al. has been deleted of five prohibited restriction sites, the region was substituted with the standard RFC[10] prefix and suffix [1].

Therefore, to establish a standardized backbone suitable for various experiments in both of these bacteria, we opted to proceed in stages. Since all the necessary restriction sites that needed to be eliminated were within a multiple cloning site, we decided to excise that portion and insert a newly synthesized fragment containing both the prefix and the suffix.

Source

The plasmid backbone was acquired from AddGene and subsequently modified

References

[1] Wang et al., 2019, Cell Chemical Biology 26, 1732–1742 December 19, 2019 ª 2019 Elsevier Ltd. https://doi.org/10.1016/j.chembiol.2019.09.003