Plasmid

Part:BBa_K510001

Designed by: Fernando Govantes   Group: iGEM11_UPO-Sevilla   (2011-09-16)
Revision as of 15:39, 16 September 2011 by Registry (Talk | contribs)

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pUC18SfiI-miniTn7BB-Cm

The miniTn7BB-Cm minitransposon cloned into SfiI-digested pUC18-Sfi. This results in the elimination of the complete pUC18-SfiI multi-cloning site, except for the duplicated SfiI sites that remain on both sides of the transposon, thus facilitating its transfer to other vectors. This plasmid can be used to maintain the mini-Tn7-Cm in E. coli and other enterics, and may be used for transposition into the genomes of non-enteric bacteria, in which it is non-replicative. The structure of the mini-Tn7BB-Cm transposon is shown in the figure below. Tn7R and Tn7L are the right and left ends of Tn7 transposon, respectively, required for recognition of the transposase machinery and transposition. FRT is the Flp recombinase target site for excision of the resistance once the transposon is inserted in the genome of a living organism. The chloramphenicol resistance cassette (Cm) is flanked by restriction sites in order to facilitate the generation of variants with different antibiotic resistance markers. The BioBrick cloning site (BCS) includes the prefix and suffix restrictions sites used in assembly standard 10, and it is the place where BioBricks are inserted for genome integration. The complete construction is flanked by SfiI restriction sites to facilitate cloning.


Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal prefix found in sequence at 4479
    Illegal suffix found in sequence at 1
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 4479
    Illegal SpeI site found at 2
    Illegal PstI site found at 16
    Illegal NotI site found at 9
    Illegal NotI site found at 4485
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 4479
    Illegal BglII site found at 3051
    Illegal BglII site found at 3322
    Illegal XhoI site found at 3408
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal prefix found in sequence at 4479
    Illegal suffix found in sequence at 2
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal prefix found in sequence at 4479
    Illegal XbaI site found at 4494
    Illegal SpeI site found at 2
    Illegal PstI site found at 16
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 1681
    Illegal SapI.rc site found at 2763


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Categories
//plasmid/chromosomalintegration
Parameters
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