Table 1.
Characteristics of sup− mutants and complementation analysis.
Figure 1.
Complementation analysis of the PQQ biosynthesis region in Ps. fluorescens strain X.
PCR fragments of this region (1–5), with different sets of genes from Ps. fluorescens X, and their ability to complement sup− mutants. Ability to complement is noted with plus (+) or minus (−). The direction of the plasposon Tn5-RL27 insertion in the derivative mutants B91, B163, A150 and ρ26 is indicated with a flag beneath the sequence. The predicted site for the unique putative promoter is also marked.
Figure 2.
Arrangement of the genes in the genomic locus of sup5 and sup6, compared to other Pseudomonas strains, and complementation analysis of the region.
The lines beneath the genomic of Ps. fluorescens X represent regions of this locus that were PCR-amplified, cloned into pBBR1MCS5 and tested for complementation. Ability to complement is noted with plus (+) or minus (−). Putative ORFs are indicated by thick coloured arrows on a line. Genes that might be organised in a putative operon are enclosed by a grey frame. The direction of the plasposon Tn5-RL27 insertion in mutants δ40 and ρ93 is indicated with a black arrow beneath the sequence (▸). Predicted sites for the unique putative promoter and operator are also marked (;). Size, genomic location and locus tag of the different ORFs sequenced in Ps. aeruginosa PA7, Ps. fluorescens Pf0–1, Ps. fluorescens SBW25, Ps. entomophila L48, Ps. syringae pv. syringae B728a,Ps. syringae pv. tomato DC3000 and Ps. syringae pv. syringae UMAF0158 are indicated.
Figure 3.
Expression levels of sup5, sup6 and orf8 in cells of Ps. fluorescens strain X and the mutants ρ26 and k36.
The expression of sup5, sup6 and orf8 was measured using rpoD as the housekeeping gene standard. Results are shown as relative expression levels compared to the expression in the wild type in LB during stationary phase. (A) Expression in the wild type grown in LB, at stationary phase; (B) Expression in the wild type grown in PDB, at stationary phase; (C) Expression in mutant k36 grown in PDB, at stationary phase; (D) Expression in mutant ρ26 grown in PDB, at stationary phase; (E) Expression in the wild type grown in LB, at exponential phase; (F) Expression in the wild type strain grown in PDB, at exponential phase. For each time point, mean values of three replicates are given; the error bars represent the standard errors of the mean.
Table 2.
Bacterial strains, plasmids, and oligonucleotides used in this study.