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

Revision as of 07:54, 29 September 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 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 BBa_J23119, a promoter that has been demonstated to be working in A. baumannii and K. pneumoniae [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

@@ Line 9: / Line 9: @@
 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]
+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===