Genomotyping of Pseudomonas putida strains using P. putida KT2440-based high-density DNA microarrays: implications for transcriptomics studies

Hendrik Ballerstedt,Rita J M Volkers,Ken N Timmis,Astrid E Mars,Jaçek Puchalka,Gerrit Eggink,Jan A M De Bont,John E Hallsworth,Jan Wery,Vitor A Martins Dos Santos,Joost Van Duuren

doi:10.1007/s00253-007-0914-z

Abstract

Pseudomonas putida KT2440 is the only fully sequenced P. putida strain. Thus, for transcriptomics and proteomics studies with other P. putida strains, the P. putida KT2440 genomic database serves as standard reference. The utility of KT2440 whole-genome, high-density oligonucleotide microarrays for transcriptomics studies of other Pseudomonas strains was investigated. To this end, microarray hybridizations were performed with genomic DNAs of subcultures of P. putida KT2440 (DSM6125), the type strain (DSM291T), plasmid pWW0-containing KT2440-derivative strain mt-2 (DSM3931), the solvent-tolerant P. putida S12, and several other Pseudomonas strains. Depending on the strain tested, 22 to 99% of all genetic elements were identified in the genomic DNAs. The efficacy of these microarrays to study cellular function was determined for all strains included in the study. The vast majority of DSM6125 genes encoding proteins of primary metabolism and genes involved in the catabolism of aromatic compounds were identified in the genomic DNA of strain S12: a prerequisite for reliable transcriptomics analyses. The genomotypic comparisons between Pseudomonas strains were used to construct highly discriminative phylogenetic relationships. DSM6125 and DSM3931 were indistinguishable and clustered together with strain S12 in a separate group, distinct from DSM291T. Pseudomonas monteilii (DSM14164) clustered well with P. putida strains.

Highlights

The sequencing and annotation of the Pseudomonas putida KT2440 genome (Nelson et al 2002) has greatly catalyzed research on this strain and other academically and biotechnologically relevant but non-sequenced P. putida strains
The genomes of S12 and other Pseudomonas strains have not been sequenced. Recent studies of their metabolic potential have been limited to comparative proteomics (Segura et al 2005; Volkers et al 2006) or transcriptomics based on the database information for P. putida KT2440
A major advantage of microarray-based comparisons of species is the ability to pinpoint differences in individual genes and intergenic regions. Through these comparisons, detailed insight was gained in the utility of P. putida KT2440-based microarrays in transcriptomics studies of different pseudomonads at the level of specific categories of biological function

Summary

Introduction

The sequencing and annotation of the Pseudomonas putida KT2440 genome (Nelson et al 2002) has greatly catalyzed research on this strain and other academically and biotechnologically relevant but non-sequenced P. putida strains. For bioprocesses involving such products, the use of solvent-tolerant P. putida strains renders advantages in terms of productivity and the application of multiphase media for product recovery (Ramos-Gonzalez et al 2003; Rojas et al 2004; Wery and De Bont 2004; Wery et al 2000; Wierckx et al 2005). Despite their biotechnological potential, the genomes of S12 and other Pseudomonas strains have not been sequenced. Recent studies of their metabolic potential have been limited to comparative proteomics (Segura et al 2005; Volkers et al 2006) or transcriptomics based on the database information for P. putida KT2440

Methods

Results

Conclusion