Part:BBa_K389011

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]