Table 1.

Oligos used in this studya

Oligo no.Oligo sequence (5′–3′)Use
744CGC GGA TCC CCA CTG GAT GCG CCG AGA TCGHybridizes withinrhaB; used to amplify recombined rhaS alleles for diagnostic PCR and sequencing
880CTA ACA TCG TCG GCA TCGHybridizes within rhaT; used to amplify recombinedrhaS alleles for sequencing
898TGA GTA AAG CTT TTA TTG CAG AAA GCC ATC CCGDownstream end ofrhaS; used to amplify rhaS alleles for chromosomal replacements
1170CCG GAA TTC TTG TGG TGA TGT GAT GCT CACUpstream of rhaS; used to amplifyrhaS alleles for chromosomal replacements
2068ATG ACC GTA TTA CAT AGT GTG GATbrhaSsequencing
2069TTA TTG CAG AAA GCC ATC CCG TCCbrhaS sequencing
2074TGG TTG CAC AGA TGG AAC AGCbrhaS sequencing
2075GTT GAG ACG TGA TGC GCT GTTbrhaS sequencing
2083GTG GGA TCC ATG ACC GTA TTA CAT AGTUpstream for diagnostic PCR on plasmid clones of all rhaS alleles
2096GCG GGA TCC GCG TTA CTC ATC TTC TTADownstream Φ(rhaT-lacZ84 and Δ133
2097CGC GAA TTC AAG GGT ATG GTT TTG CAGUpstream Φ(rhaT-lacZ133
2130GGC CTG GCT GGC AGA CCA TTT TGSDM; RhaS E181Ala
2131CAT CGG CAA AAT GGT CTGDiagnostic PCR; RhaS E181Ala
2134CCA TTT TGC CGC AGA GGT GAA TTGSDM; RhaS E186Ala
2135CAT CCC AAT TCA CCT CTGDiagnostic PCR; RhaS E186Ala
2136TTG CCG ATGCAG TGA ATT GGSDM; RhaS E187Ala
2137CGG CAT CCC AAT TCA CTGDiagnostic PCR; RhaS E187Ala
2138CCG TGG CGGCAC AAT TTT CTSDM; RhaS D195Ala
2139CGC AGT GAA AGA GAA AAT TGT GDiagnostic PCR; RhaS D195Ala
2141TGT CAG TAA CGC TGG CTGDiagnostic PCR; RhaS E236Ala
2142CGT TAC TGC AAT CGC CTA TCSDM; RhaS D241Ala
2143CAC AGC GAT AGG CGA TTGDiagnostic PCR; RhaS D241Ala
2146TCA CCG CGT GCA ATT CGC CASDM; RhaS D268Ala
2147CCG TCC CTG GCG AAT TGDiagnostic PCR; RhaS D268Ala
2148AGG GAC GGGCAG GCT TTC TSDM; RhaS D274Ala
2149TTA TTG CAG AAA GCC TGDiagnostic PCR; RhaS D274Ala
2152CCG GAA TTC ACT TAA TGC CGT GAT TGUpstream Φ(rhaT-lacZ84
2154TGG CTG GAG GCT CAT TTT GCCSDM; RhaS D182Ala
2155ACG CCA CAG CGC AGC CAG CGT TASDM; RhaS E236Ala
2156CCT CAT CGG CAA AAT GAGDiagnostic PCR; RhaS D182Ala
2161TTT GTT TGC GTT TAC TGG CAG ATADownstream Plac-bet exo kan
2162ACG GCA ACG GCC TTG AAC TGA AATUpstream Plac-bet exo kan
2185TTC GCC GAGCAT TTA ACT GGT CSDM; RhaS E261Ala
2186GCG GTG ACC AGT TAA ATGDiagnostic PCR; RhaS E261Ala
  • a Oligos were used for cloning, diagnostic or regular PCR, and site-directed mutagenesis (SDM). Regions not complementary to wild-type rha genes are underlined.

  • b Oligos IRD41 dye labeled for use in a LI-COR automated sequencer.