Genome-wide transcription profiling of Corynebacterium glutamicum after heat shock and during growth on acetate and glucose

• Published in: Journal of biotechnology 2002-09, Vol.98 (2), p.255-268
• Main Authors: Muffler, Andrea, Bettermann, Sandra, Haushalter, Michael, Hörlein, Andreas, Neveling, Ute, Schramm, Maren, Sorgenfrei, Oliver
• Format: Article
• Language: English
• Subjects: Acetate metabolism; Acetates - metabolism; Biological and medical sciences; Biology of microorganisms of confirmed or potential industrial interest; Biotechnology; Carbon - metabolism; Corynebacterium - genetics; Corynebacterium - physiology; Corynebacterium glutamicum; Culture Media - metabolism; Fundamental and applied biological sciences. Psychology; Gene Expression; Gene Expression Profiling - methods; Gene Expression Regulation; Genome, Bacterial; Glucose - metabolism; Heat shock; Heat-Shock Response - genetics; Heat-Shock Response - physiology; Mission oriented research; Models, Chemical; Oligonucleotide Array Sequence Analysis - instrumentation; Oligonucleotide Array Sequence Analysis - methods; ORF-based DNA-array; Physiology and metabolism; Polymerase Chain Reaction; Sensitivity and Specificity; Sequence Analysis, DNA - methods; Shotgun DNA-array
• ISSN: 0168-1656; 1873-4863
• DOI: 10.1016/S0168-1656(02)00136-0

To monitor the global gene expression of Corynebacterium glutamicum we established two formats of DNA-arrays on nylon membranes. We produced an ordered DNA-array of PCR fragments from a shotgun library of C. glutamicum representing a threefold coverage of the genome. With this format we studied genome-wide transcriptional changes after heat shock. Sequence and subsequent BLAST analysis of PCR fragments with elevated expression after heat shock revealed PCR fragments harboring genes that encode several proteins of the heat shock family, proteins of the oxidative stress response and proteins with unknown function. DNA-arrays based on PCR fragments representing 2804 annotated ORFs of C. glutamicum were used to monitor the transcript levels during growth on acetate and glucose. We determined minimal detectable ratios and compared labeling approaches with random hexamers and ORF-specific primers. ORF-based DNA-array analysis with different labeling approaches showed similar results: e.g. increased mRNA levels of the pta- ack operon, aceA, aceB and genes encoding phosphoenolpyruvate carboxykinase and enzymes of the citric acid cycle during growth on acetate and elevated mRNA levels of some enzymes of the glycolytic pathway and lactate dehydrogenase upon growth on glucose. These results demonstrate that shotgun DNA-arrays and ORF-based DNA-arrays are appropriate tools to study physiology of microorganism.