Part:BBa_J153000:Design

Broad-host-range shuttle vector pPMQAK1

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  INCOMPATIBLE WITH RFC[12]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 7676
  Illegal SpeI site found at 2
  Illegal PstI site found at 16
  Illegal NotI site found at 9
  Illegal NotI site found at 7682
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 7676
  Illegal XhoI site found at 6569
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal prefix found at 7676
  Illegal suffix found at 2
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal prefix found at 7676
  Plasmid lacks a suffix.
  Illegal XbaI site found at 7691
  Illegal SpeI site found at 2
  Illegal PstI site found at 16
  Illegal AgeI site found at 1061
  Illegal AgeI site found at 1462
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal BsaI.rc site found at 6715
  

Design Notes

As cloning by plasmid preparation and subsequent digestion/ligation was problematic due to low copy number and difficulties in digesting the RSF1010-derived part from pAWG1.1, pPMQAK1 was made using PCR and subsequent digestion/ligation.

Source

pPMQAK1 was constructed by ligating a PCR product from the pAWG1.1 plasmid, corresponding to the RSF1010 replicon, with a PCR product from the pSB1AK3 plasmid, corresponding to the BioBrick cloning site with flanking functional sequences plus the ampicillin and kanamycin resistance cassettes.

References

Huang, H.H., Camsund, D., Lindblad, P. and Heidorn, T. (2010) Design and characterization of molecular tools for a Synthetic Biology approach towards developing cyanobacterial biotechnology. Nucleic Acids Res, 38, 2577-2593.