title: Construction and validation of a Sinorhizobium meliloti whole genome DNA microarray: genome-wide profiling of osmoadaptive gene expression.
abstract: Based on the complete Sinorhizobium meliloti genome sequence we established DNA microarrays as a comprehensive tool for systematic genome-wide gene expression analysis in S. meliloti 1021. For these PCR fragment-based microarrays, called Sm6kPCR, a collection of probes for the 6207 predicted protein-coding genes consisting of 6046 gene-specific PCR fragments and 161 70 mer oligonucleotides was arrayed in high density on glass slides. To obtain these PCR fragments primer pairs were designed to amplify internal gene-specific DNA fragments of 80-350 bp. Additionally, these primers were characterized by a 5' extension that allowed for reamplification using standard primers after the first amplification employing the specific primers. In order to ascertain the quality of the Sm6kPCR microarrays and to validate gene expression studies in S. meliloti parallel hybridizations based on RNA samples obtained from cells cultured under identical conditions were performed. In addition, gene expression in S. meliloti in response to an osmotic upshift imposed by the addition of 0.38 M NaCl was monitored. 137 genes were identified showing significant changes in gene expression resulting from the osmotic upshift. From these genes 52 were induced and 85 genes were repressed. Among the genes displaying different RNA levels some functional groups could be identified that are particularly remarkable. Repression was observed for 8 genes related to motility and chemotaxis, 7 genes encoding amino acid biosynthesis enzymes and 15 genes involved in iron uptake whereas 14 genes involved in transport of small molecules and 4 genes related to polysaccharide biosynthesis were induced.
authors.affiliation: Lehrstuhl für Genetik, Universität Bielefeld, Universitätsstrasse 25, D-33615 Bielefeld, Germany.
authors.forename: Silvia
authors.initials: S
authors.lastname: Rüberg
authors.forename: Zhe-Xian
authors.initials: ZX
authors.lastname: Tian
authors.forename: Elizaveta
authors.initials: E
authors.lastname: Krol
authors.forename: Burkhard
authors.initials: B
authors.lastname: Linke
authors.forename: Folker
authors.initials: F
authors.lastname: Meyer
authors.forename: Yiping
authors.initials: Y
authors.lastname: Wang
authors.forename: Alfred
authors.initials: A
authors.lastname: Pühler
authors.forename: Stefan
authors.initials: S
authors.lastname: Weidner
authors.forename: Anke
authors.initials: A
authors.lastname: Becker
mesh_terms: D002874:Chromosome Mapping / Q000295:instrumentation* / Q000379:methods; D004867:Equipment Design; D019544:Equipment Failure Analysis; D020869:Gene Expression Profiling / Q000295:instrumentation* / Q000379:methods; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology*; D016680:Genome, Bacterial; D020411:Oligonucleotide Array Sequence Analysis / Q000295:instrumentation* / Q000379:methods; D016133:Polymerase Chain Reaction / Q000295:instrumentation / Q000379:methods; D016962:Sinorhizobium meliloti / Q000145:classification / Q000235:genetics*; D013045:Species Specificity; D014882:Water-Electrolyte Balance / Q000235:genetics*