Part:BBa_K606003

Multi-host replicative plasmid, for B. Subtilis and E. Coli

This plasmid is an multihost plasmid, replicated both in E. Coli at high copy number (>200) and low copy in B. Subtilis (about 20).

This plasmid is ideal for importing parts in Subtilis: all the cloning can be done in E. Coli, as well as the amplification. Once the plasmid is extracted, you can incorporate it directly in B. Subtilis using electroporation.

This plasmid is Ampicilyn resistant in E. Coli, and is tetracycline resistant in both B. Subtilis and E. Coli.

The biobrick BBa_J04450 is inserted between the E X and SP site, as a negative cloning control in E. Coli

Beware, there is a pT7 promoter hided in the original sequence of one of the plasmid. This plasmid may be incompatible if you are working in a system with T7 polymerase.

This plasmid was properly characterized. You can find out all the remarks and comments on our wiki at this page [http://2011.igem.org/Team:Paris_Bettencourt/MultiHost].

Usage and Biology

Move your constructions from E. Coli to B. Subtilis, and express your proteins. We encourage you tu use this plasmid in combination with the pVeg promoter (BBa_K143012) and SpoVg RBS (K143021)

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 17
  Illegal XbaI site found at 2
  Illegal SpeI site found at 5918
  Illegal PstI site found at 5904
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 17
  Illegal SpeI site found at 5918
  Illegal PstI site found at 5904
  Illegal NotI site found at 9
  Illegal NotI site found at 5909
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 17
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 17
  Illegal XbaI site found at 2
  Illegal SpeI site found at 5918
  Illegal PstI site found at 5904
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 17
  Illegal XbaI site found at 2
  Illegal SpeI site found at 5918
  Illegal PstI site found at 5904
  Illegal NgoMIV site found at 3573
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal BsaI site found at 2560
  Illegal BsaI site found at 4550
  Illegal SapI.rc site found at 5632