Counter-selectable markers

Desired Characteristics

1. Short sequence
   • shorter markers will enable cheaper synthesis of codon variants with different stabilities.
2. Single gene responsible for selection
   • enables easier determination of mutations responsible for phenotype
   • for instance, a system where a constitutive repressor turned off a toxin could accumulate mutations in either the repressor or the toxin
3. Strong selective pressure
   • less false positives
4. On the chromosome?
   • Not sure how mutations on a plasmid differ...

Candidates

gene(selection condition)

1. lacY (t-o-nitrophenyl-B-D-galactopyranoside)
2. thyA (trimethoprim)
3. pheS (p-chlorophenylalanine)
   • Relaxed substrate specificity mutant of phenylalanine tRNA transferase that will incorporate p-chlorophenylalanine into proteins, killing the cell
4. ?(phage)
   • probably >1 gene can confer resistance?
   • previous work (Novick/Szilard) has mapped mutation rates confering T1 resistance in chemostats.
5. pyrF (5FoA)
   • Jeff has ordered synthesized/BB'd
   • in pyrF- strains can be selected on medium w/o Uracil, so can be selected and coutner-selected.
6. sacB
7. tet?
8. others???