Genome-wide transcription profiling of Corynebacterium glutamicum after heat shock and during growth on acetate and glucose.
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.
Axaron Bioscience AG, Im Neuenheimer Feld 515, 69120 Heidelberg, Germany.
Andrea
A
Muffler
Sandra
S
Bettermann
Michael
M
Haushalter
Andreas
A
Hörlein
Ute
U
Neveling
Maren
M
Schramm
Oliver
O
Sorgenfrei
D000085:Acetates / Q000378:metabolism; D002244:Carbon / Q000378:metabolism; D003352:Corynebacterium / Q000235:genetics / Q000502:physiology*; D003470:Culture Media / Q000378:metabolism; D015870:Gene Expression; D020869:Gene Expression Profiling / Q000379:methods*; D005786:Gene Expression Regulation; D016680:Genome, Bacterial; D005947:Glucose / Q000378:metabolism; D018869:Heat-Shock Response / Q000235:genetics / Q000502:physiology*; D008956:Models, Chemical; D020411:Oligonucleotide Array Sequence Analysis / Q000295:instrumentation / Q000379:methods; D016133:Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA / Q000379:methods
D000085:Acetates; D003470:Culture Media; D002244:Carbon; D005947:Glucose