Part:BBa_K1321330

TetR repressed RFP in pSEVA331-Bb

This is a composite part containing the TetR gene generator and an RFP generator behind pTet promoter (BBa_K577895) in pSEVA331-Bb backbone (part BBa_K1321300). This construct is a member of the G.xylinus genetic engineering toolbox. This provides both the TetR generator (tetR is absent from G. xylinus genome) and the pTet promoter for inducible expression in G.xylinus.BBa_K1321330 is a member of the G.xylinus genetic engineering toolkit (parts BBa_K1321295 - BBa_K1321332).

G.xylinus toolkit was designed to ease genetic engineering of cellulose-based biomaterials using the cellulose synthesizing bacterium Gluconacetobacter xylinus (parts Bba_K1321305 and BBa_K1321306). As no registry parts had been tested in G.xylinus, the aim of this toolkit was to determine the parts usable in G.xylinus and to characterize them in this host. pSEVA331-Bb is a non-standard broad host range plasmid capable of replication in G.xylinus and E.coli (a shuttle vector) and was selected because the registry's standard plasmid backbone pSB1C3 can not be used for G.xylinus engineering.

NOTE: Because the registry's standard plasmid backbone pSB1C3 is not capable of replication in Gluconacetobacter species, the G.xylinus genetic engineering toolkit is housed mainly in pSEVA331-Bb. pSEVA331-Bb is a non-standard backbone, which therefore can't be quality controlled by and maintained in the Registry. However, in order to make the G.xylinus toolkit available for the synthetic biology community, Imperial iGEM 2014 team has made it freely available upon request, with quality control provided (see Experience). To request, please contact Imperial iGEM 2014 team.

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal prefix found in sequence at 4697
  Illegal suffix found in sequence at 1826
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal EcoRI site found at 4697
  Illegal SpeI site found at 1827
  Illegal PstI site found at 1841
  Illegal NotI site found at 1834
  Illegal NotI site found at 4703
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal EcoRI site found at 4697
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal prefix found in sequence at 4697
  Illegal suffix found in sequence at 1827
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal prefix found in sequence at 4697
  Illegal XbaI site found at 4712
  Illegal SpeI site found at 1827
  Illegal PstI site found at 1841
  Illegal NgoMIV site found at 3016
  Illegal AgeI site found at 1529
  Illegal AgeI site found at 1641
  
• 1000
  COMPATIBLE WITH RFC[1000]