Verification that the nonsense mutation in citS leads to a defect in the citrate import pathway.
A) Sanger sequencing verification of the Solexa citS sequence. Sequences are shown in the following order: the published reference sequence first, followed by the Solexa results for the citS gene in strains 168 and JH642, and the Sanger results for both strains shown last. B, C) Rescue of the citS- phenotype of JH642. Strains were plated to obtain single colonies on minimal media supplemented with glucose (B) or citrate (C) as the sole carbon source. The strains shown are: the prototroph SMY, the lab strain JH642, a JH642-derived strain that is ‘cured’ of its growth-dependence on the amino acids tryptophan and phenylalanine (JH642 trp+, phe+), and finally, the latter strain, transformed with the complete citS gene from SMY (JH642 trp+ phe+ citS+). JH642 trp+ phe+ is able to utilize glucose but not citrate. JH642 trp+ phe+ citS+ is able to utilize both glucose and citrate.