Composite

Part:BBa_K389011

Designed by: Jonas Aretz   Group: iGEM10_Bielefeld-Germany   (2010-08-10)

VirA screening device

This part contains:

  • a mutated virG response regulator from the VirA/G receptor system from Agrobacterium tumefaciens which works in Escherichia coli without an rpoA gene from A. tumefaciens
  • a kanamycin resistance under the control of a vir promoter


Usage and Biology

This device is used to screen virA receptor mutants. The virA receptor gene is mutated in the BBa_K389010 BioBrick. The better the receptor detects a substance, the more VirG is phosphorylated and the stronger the kanamycin resistance from this BioBrick is expressed. The screening system for a mutated virA gene contains this BioBrick in a plasmid with R6K ori and the BioBrick BBa_K389010 in a plasmid with ColE1 ori. Both plasmids are transformed to and screened in E. coli EC100D because plasmids with R6K ori are only replicated in pir+ or pir116 E. coli strains. Once a receptor with high sensitivity for a screened substance is found, the plasmids are isolated and transformed to e.g. E. coli TOP10. Because the R6K ori does not work in this strain because it is not a pir+ / pir116 strain, it is easy to separate the mutated virA BioBrick from the screening plasmid.


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 7
    Illegal NheI site found at 30
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


[edit]
Categories
//cds/selectionmarker/antibioticresistance
//cds/transcriptionalregulator/activator
//chassis/prokaryote/ecoli
//classic/measurement/uncategorized
Parameters
None