Plasmid_Backbone

Part:BBa_K4166006

Designed by: George Amoako   Group: iGEM22_Alma   (2022-10-04)


camps +8 -11

Runaway plasmid replication (+8) Preventing R-loop formation (-11)

Plasmid Copy Number and Expression - Alma 2022

We created a series of mutants of BBa_J04450 in the origin of replication, and wanted to both quantify the copy number and see the effect the copy number has on gene expression.

We isolated both genomic DNA and plasmid DNA from the cells and ran a serial dilution of these on a gel to observe the following.

T--Alma--gel 006.png
T--Alma--pcnPlate.png

From the intensity of these bands we estimate the copy number of this origin at ~163 copies/cell. We found that this combination of mutations lowers copy number, and also lowers expression (as expected).

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Plasmid lacks a prefix.
    Plasmid lacks a suffix.
  • 12
    INCOMPATIBLE WITH RFC[12]
    Plasmid lacks a prefix.
    Plasmid lacks a suffix.
  • 21
    INCOMPATIBLE WITH RFC[21]
    Plasmid lacks a prefix.
    Plasmid lacks a suffix.
  • 23
    INCOMPATIBLE WITH RFC[23]
    Plasmid lacks a prefix.
    Plasmid lacks a suffix.
  • 25
    INCOMPATIBLE WITH RFC[25]
    Plasmid lacks a prefix.
    Plasmid lacks a suffix.
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Plasmid lacks a prefix.
    Plasmid lacks a suffix.


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