Part:BBa_K917010
Kanamycin resistance+lac promoter+RFP+sacB selection-counterselection cassette
This is an improved version of parts BBa_K322922 and K322923, 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, together with a kanamycin (BBa_P1003) or chloramphenicol (BBa_K322210) resistance marker. 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
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 1756
Illegal AgeI site found at 1868 - 1000COMPATIBLE WITH RFC[1000]
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