Plasmid_Backbone

Part:pSB3C02

Designed by: Vinoo Selvarajah   Group: Registry   (2019-04-04)


pEven2 Loop Vector based on pSB3C5

pEven2 Loop Vector based on pSB3C5


iGEM Type IIS Vectors

pEven Loop Vectors (for Level 2)

When four Level 1 parts (TUs) are assembled into a Multi-Transcriptional Unit (MTU) into the following plasmid backbones (with BBa_J04455), the MTU will be flanked by BsaI restriction sites and these 4 bp Fusion Sites.

Registry Name Loop Name Fusion Site 5' MTU Fusion Site 3'
pSB3C01 pEven1 GGAG MTU 1 TACT
pSB3C02 pEven2 TACT MTU 2 AATG
pSB3C03 pEven3 AATG MTU 3 GCTT
pSB3C04 pEven4 GCTT MTU 4 CGCT

Note:

  • The documented sequence shows an additional BsaI site. This would impact assembly out of (but not into) this set of vectors. A new set of vectors should be created or used in the long term.
  • Currently, Registry samples of these plasmid backbones have not been fully sequenced. Only their prefix and suffix along with their insert BBa_J04455 have been verified.


The iGEM Type IIS assembly standard is based on MoClo and Loop


Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Plasmid lacks a prefix.
    Plasmid lacks a suffix.
    Illegal EcoRI site found at 2713
    Illegal PstI site found at 12
  • 12
    INCOMPATIBLE WITH RFC[12]
    Plasmid lacks a prefix.
    Plasmid lacks a suffix.
    Illegal EcoRI site found at 2713
    Illegal NheI site found at 1399
    Illegal PstI site found at 12
  • 21
    INCOMPATIBLE WITH RFC[21]
    Plasmid lacks a prefix.
    Plasmid lacks a suffix.
    Illegal EcoRI site found at 2713
  • 23
    INCOMPATIBLE WITH RFC[23]
    Plasmid lacks a prefix.
    Plasmid lacks a suffix.
    Illegal EcoRI site found at 2713
    Illegal PstI site found at 12
  • 25
    INCOMPATIBLE WITH RFC[25]
    Plasmid lacks a prefix.
    Plasmid lacks a suffix.
    Illegal EcoRI site found at 2713
    Illegal PstI site found at 12
    Illegal AgeI site found at 990
    Illegal AgeI site found at 1313
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Plasmid lacks a prefix.
    Plasmid lacks a suffix.
    Illegal BsaI site found at 285
    Illegal BsaI site found at 2719
    Illegal BsaI.rc site found at 6


Uppsala 2020's Improvement

TEAMNAME UofUppsala YEAR 2020 Authors Nuria Garriga Alonso, Bjorn Ancker Persson, Tereza Hubackova, Dorottya Marko, Hui Yu

The illegal BsaI site was removed by PCR mutagenesis to obtain the new pEven2 plasmid pSB3C12 (BBa_K3425002). This new plasmid was used for various clonings and it works as expected.

Moreover, pSB3C01 was also used for cloning and it also works as expected. However, we recommend using the new plasmids without an illegal BsaI site in order to avoid compromising the efficiency.

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