Composite

Part:BBa_K917002

Designed by: Elitsa Peeva   Group: iGEM12_Edinburgh   (2012-09-17)
Revision as of 20:51, 1 October 2012 by Pandeli (Talk | contribs)

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lac promoter plus RFP and sacB

This is an improved version of part BBa_K322921 (French & Kowal, 2010), which consisted of the Bacillus subtlis 168 levansucrase gene, sacB, a widely used counterselectable marker which is lethal when expressed in E. coli and other Gram negative bacteria in the presence of high sucrose concentrations (Gay, Coq, Strinmetz, Ferrari, & Hoch, 1983). In this version, the lac promoter and Red Fluorescent Protein (RFP) from BBa_J04450 (but without the terminator) have been added. The provides a controllable promoter for sacB and also a marker gene. This is valuable as sacB counter-selection is notoriously prone to escape due to loss-of-function mutations in sacB. By including RFP, we can distinguish true loss of the cassette (giving white colonies) from loss of sacB function (which should still give red colonies). This will make our experiments much easier to interpret.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 781
    Illegal AgeI site found at 893
  • 1000
    COMPATIBLE WITH RFC[1000]


[edit]
Categories
Parameters
None