title: Pseudomonas putida KT2440 genome update by cDNA sequencing and microarray transcriptomics.
abstract: Pseudomonas putida KT2440 is a completely sequenced biosafety strain that has retained its capability to survive and function in the environment. The global mRNA expression profiles of the KT2440 strain grown at 10°C and 30°C were determined by deep cDNA sequencing to refine the genome annotation. Transcriptome sequencing identified 36 yet unknown small non-coding RNAs, 143 novel ORFs in 106 intergenic regions, 42 unclassified genes and eight highly expressed leaderless mRNA transcripts. The genome coordinates of eight genes and the organization of 57 operons were corrected. No overrepresented sequence motifs were detected in the 5'-untranslated regions. The 50 most highly expressed genes made up 60% of the total mRNA pool. Comparison of cDNA sequencing, Affymetrix and Progenika microarray data from the same mRNA preparation revealed a higher sensitivity and specificity of cDNA sequencing, a relatively poor correlation between the normalized cDNA reads and microarray signal intensities, and a systematic signal-dependent bias of microarrays in the detection of differentially regulated genes. The study demonstrates the power of next-generation cDNA sequencing for the quantitation of mRNA transcripts and the refinement of bacterial genome annotation.
authors.affiliation: Klinische Forschergruppe, Medizinische Hochschule Hannover, Carl-Neuberg-Strasse 1, Hannover, Germany.
authors.forename: Sarah
authors.initials: S
authors.lastname: Frank
authors.forename: Jens
authors.initials: J
authors.lastname: Klockgether
authors.forename: Petra
authors.initials: P
authors.lastname: Hagendorf
authors.forename: Robert
authors.initials: R
authors.lastname: Geffers
authors.forename: Ulrike
authors.initials: U
authors.lastname: Schöck
authors.forename: Thomas
authors.initials: T
authors.lastname: Pohl
authors.forename: Colin F
authors.initials: CF
authors.lastname: Davenport
authors.forename: Burkhard
authors.initials: B
authors.lastname: Tümmler
mesh_terms: D004269:DNA, Bacterial / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D020869:Gene Expression Profiling; D016680:Genome, Bacterial; D058977:Molecular Sequence Annotation; D020411:Oligonucleotide Array Sequence Analysis; D016366:Open Reading Frames; D016958:Pseudomonas putida / Q000235:genetics*; D058727:RNA, Small Untranslated / Q000235:genetics; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA / Q000379:methods; D020506:Untranslated Regions
chemical_list: D004269:DNA, Bacterial; D018076:DNA, Complementary; D058727:RNA, Small Untranslated; D020506:Untranslated Regions