Coding

Part:BBa_K3888005

Designed by: Linhan Lei   Group: iGEM21_SJTang   (2021-10-01)
Revision as of 18:52, 20 October 2021 by RyanLei (Talk | contribs)

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UppE Upstream Flanking Region

UppE Upstream Flanking Region
This basic part is the upstream flanking region of UppE for R. palustris Together with UppE downstream flanking region(BBa_K3888006), it forms UppE Knockout(BBa_K3888998).
Upp encodes part of the unipolar polysaccharide (UPP) and it has been proved that UPP is important for adhesion when light and organic substrates are used for growth for R. palustris UppE(RPA2750) is located on 3120549..3122105 on the complete genome of R. palustris. Deletion Plasmid construction.png

Figure 1: Illustration of the general form of plasmid used to create marked knockout strains of R. palustris.

The origin of replication (ori), origin of transfer (oriT) and sacB (blue) are part of the delivery plasmid. The green sequence is inserted into the multiple cloning site of the delivery plasmid. The left and right fragments (green) are sequences amplified from the R. palustris genome and used in overlap extension PCR to generate a recombinant product containing (a deletion and a restriction site）. The restriction site is used to introduce the resistance cassette (purple). This interrupts the gene even if no deletion is present, and allows selection of successful transformants.
According to this model, we constructed UppE knockout plasmid below. UppE Deletion Plasmid Map.png

Figure 2: Deletion plasmid of UppE constructed according to the method mentioned above. UppE Upstream Flanking region is located from 1021bp to 2056bp.

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 694
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 694
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 694
    Illegal BglII site found at 333
    Illegal XhoI site found at 826
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 694
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 694
    Illegal NgoMIV site found at 38
    Illegal NgoMIV site found at 119
    Illegal NgoMIV site found at 157
    Illegal NgoMIV site found at 831
    Illegal NgoMIV site found at 943
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 538


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