|
| Status |
Public on Sep 05, 2011 |
| Title |
wildtype: 21.25 µM copper_vs_1.25 µM copper rep2 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
wt; 21.25 µM copper
|
| Organism |
Corynebacterium glutamicum ATCC 13032 |
| Characteristics |
treatment: 21.25 uM copper
genotype: wild type
|
| Treatment protocol |
see growth protocol
|
| Growth protocol |
Bacterial strains, media and growth conditions. For growth experiments or DNA microarray analysis 5 ml of brain heart infusion (BHI) medium (Difco Laboratories, Detroit, MI) were inoculated with colonies from a fresh BHIS agar plate (BHI agar with 0.5 M sorbitol) and incubated for 6 hours at 30°C and 170 rpm. This first preculture was used to inoculate a 500-ml shake flask containing 50 ml CGXII minimal medium (19) with 4% (w/v) glucose added as carbon source with 1.25 µM or 21.25 µM CuSO4. The second preculture was cultivated overnight at 30°C and then used to inoculate the main culture to an optical density at 600 nm (OD600) of ~1 in a 500-ml shake flask containing 50 ml CGXII minimal medium (19) with 4% (w/v) glucose with 1.25 µM or 21.25 µM CuSO4. For DNA microarray analysis cells were harvested at an OD600 of 5-6.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Preparation of total RNA were carried out as described in Lange, C., D. Rittmann, V. F. Wendisch, M. Bott, and H. Sahm. 2003. Global expression profiling and physiological characterization of Corynebacterium glutamicum grown in the presence of L-valine. Appl. Environm. Microbiol. 69:2521-2532
|
| Label |
Cy3
|
| Label protocol |
Synthesis of fluorescently labelled cDNA were carried out as described in Lange, C., D. Rittmann, V. F. Wendisch, M. Bott, and H. Sahm. 2003. Global expression profiling and physiological characterization of Corynebacterium glutamicum grown in the presence of L-valine. Appl. Environm. Microbiol. 69:2521-2532
|
| |
|
| Channel 2 |
| Source name |
wt; 1.25 µM copper
|
| Organism |
Corynebacterium glutamicum ATCC 13032 |
| Characteristics |
treatment: 1.25 µM copper
genotype: wild type
|
| Treatment protocol |
see growth protocol
|
| Growth protocol |
Bacterial strains, media and growth conditions. For growth experiments or DNA microarray analysis 5 ml of brain heart infusion (BHI) medium (Difco Laboratories, Detroit, MI) were inoculated with colonies from a fresh BHIS agar plate (BHI agar with 0.5 M sorbitol) and incubated for 6 hours at 30°C and 170 rpm. This first preculture was used to inoculate a 500-ml shake flask containing 50 ml CGXII minimal medium (19) with 4% (w/v) glucose added as carbon source with 1.25 µM or 21.25 µM CuSO4. The second preculture was cultivated overnight at 30°C and then used to inoculate the main culture to an optical density at 600 nm (OD600) of ~1 in a 500-ml shake flask containing 50 ml CGXII minimal medium (19) with 4% (w/v) glucose with 1.25 µM or 21.25 µM CuSO4. For DNA microarray analysis cells were harvested at an OD600 of 5-6.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Preparation of total RNA were carried out as described in Lange, C., D. Rittmann, V. F. Wendisch, M. Bott, and H. Sahm. 2003. Global expression profiling and physiological characterization of Corynebacterium glutamicum grown in the presence of L-valine. Appl. Environm. Microbiol. 69:2521-2532
|
| Label |
Cy5
|
| Label protocol |
Synthesis of fluorescently labelled cDNA were carried out as described in Lange, C., D. Rittmann, V. F. Wendisch, M. Bott, and H. Sahm. 2003. Global expression profiling and physiological characterization of Corynebacterium glutamicum grown in the presence of L-valine. Appl. Environm. Microbiol. 69:2521-2532
|
| |
|
| |
| Hybridization protocol |
Custom-made whole-genome DNA microarrays for C. glutamicum ATCC 13032 printed with 70-mer oligonucleotides were obtained from Operon (Cologne, Germany) and are based on the genome sequence entry NC_006958. Hybridisation and stringent washing of the microarrays were performed according to the instructions of the supplier. Hybridisation was carried out for 16 - 18 h at 42°C using a MAUI hybridization system (BioMicro Systems, Salt Lake City, USA).
|
| Scan protocol |
After washing the microarrays were dried by centrifugation (5 min, 1600 g) and fluorescence was determined at 532 nm (Cy3-dUTP) and 635 nm (Cy5-dUTP) with 10 µm resolution using an Axon GenePix 6000 laser scanner (Axon Instruments, Sunnyvale, U.S.A).
|
| Data processing |
Quantitative image analysis was carried out using GenePix image analysis software and results were saved as GPR-file (GenePix Pro 6.0, Axon Instruments). For data normalization, GPR-files were processed using the BioConductor/R-packages limma and marray (www.bioconductor.org). Processed and normalized data as well as experimental details (according to MIAME) were stored in the in-house microarray database for further analysis.
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| |
|
| Submission date |
Feb 24, 2011 |
| Last update date |
Apr 14, 2021 |
| Contact name |
Tino Polen |
| E-mail(s) |
t.polen@fz-juelich.de
|
| Organization name |
Forschungszentrum Jülich GmbH
|
| Department |
IBG-1: Biotechnology
|
| Street address |
Leo Brandt Str.
|
| City |
Juelich |
| State/province |
NRW |
| ZIP/Postal code |
52425 |
| Country |
Germany |
| |
|
| Platform ID |
GPL9860 |
| Series (1) |
| GSE27510 |
The Two-component Signal Transduction System CopRS of Corynebacterium glutamicum is Required for Adaptation to Copper-excess Stress |
|
| Relations |
| Reanalyzed by |
GSM5197179 |
