|
| Status |
Public on Apr 01, 2013 |
| Title |
Mixed culture type 2_mono culture type 2 [batch cultures] |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
Lbul/Scer
|
| Organisms |
Lactobacillus delbrueckii subsp. bulgaricus; Saccharomyces cerevisiae |
| Characteristics |
culture type: Combined mono cultures of L. bulgaricus and S. cerevisiae
cultivation type: Batch
experiment: Reference to test cross-hybridisation
|
| Treatment protocol |
Sampling for transcriptome analysis from pure and mixed cultures was performed as previously described, using liquid nitrogen for rapid quenching of metabolism (J.Biol.Chem. 283:23524-23532). Samples were stored at -80°C in a mixture of phenol/chloroform and TEA buffer until further processing for total RNA extraction.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
L. bulgaricus total RNA from pure and mixed cultures was isolated as previously described (Appl.Environ.Microbiol. 76:7775-7784). 500 mg of 0.1 mm zirconium beads (Biospec Products Inc., OK, USA) were added to the extraction tubes to ensure efficient mechanical lysis of cells.
|
| Label |
Cy5
|
| Label protocol |
Processing of RNA, labeling and hybridization were performed as in Appl.Environ.Microbiol. 76:7775-7784. Each sample was split in two after purification and the halves were labeled with cyanine3 and cyanine 5.
|
| |
|
| Channel 2 |
| Source name |
Lbul mono
|
| Organism |
Lactobacillus delbrueckii subsp. bulgaricus |
| Characteristics |
culture type: Mono culture
cultivation type: Batch
experiment: Reference to test cross-hybridisation
|
| Treatment protocol |
Sampling for transcriptome analysis from pure and mixed cultures was performed as previously described, using liquid nitrogen for rapid quenching of metabolism (J.Biol.Chem. 283:23524-23532). Samples were stored at -80°C in a mixture of phenol/chloroform and TEA buffer until further processing for total RNA extraction.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
L. bulgaricus total RNA from pure and mixed cultures was isolated as previously described (Appl.Environ.Microbiol. 76:7775-7784). 500 mg of 0.1 mm zirconium beads (Biospec Products Inc., OK, USA) were added to the extraction tubes to ensure efficient mechanical lysis of cells.
|
| Label |
Cy3
|
| Label protocol |
Processing of RNA, labeling and hybridization were performed as in Appl.Environ.Microbiol. 76:7775-7784. Each sample was split in two after purification and the halves were labeled with cyanine3 and cyanine 5.
|
| |
|
| |
| Hybridization protocol |
For each array, 0.3 µg of cDNA labeled with either Cyanine 3 or Cyanine 5 was hybridized using the solutions and following the protocol delivered by Agilent (version 5.5) for 8x15K slides.
|
| Scan protocol |
Slides were scanned using an Agilent microarray scanner (G2565BA). Lasers of wavelengths at 532 and 635 nm were used to excite Cyanine3 and Cyanine5 dyes, respectively. Fluorescent images were captured as multi-image-tagged image file format and analyzed with Imagene software (Axon) (BioDiscovery, Marina del Rey, USA).
|
| Description |
Lbul/Scer over Lbul mono
Mono culture L. bulgaricus RNA mixed with mono culture S. cerevisiae RNA compared to mono culture L. bulgaricus RNA (batch cultures)
|
| Data processing |
The extent of hybridization was derived from a median value of pixel-by-pixel ratios. Spots were normalized using Lowess. Differential regulation was determined by false-discovery rate (FDR) from the Cyber-T p-values by means of multiple testing connections. Differential regulation was defined as a two-fold or higher differential expression with a FDR cut-off value of 0.05 or lower. Biological replicates were pooled in the analysis.
|
| |
|
| Submission date |
Apr 01, 2013 |
| Last update date |
Apr 01, 2013 |
| Contact name |
Sander Sieuwerts |
| Organization name |
Arla Foods
|
| Department |
Arla Scientific Innovation Centre
|
| Lab |
Microbial Cultures and Enzymes
|
| Street address |
rørdrumvej 2
|
| City |
Brabrand |
| ZIP/Postal code |
8220 |
| Country |
Denmark |
| |
|
| Platform ID |
GPL16855 |
| Series (1) |
| GSE45623 |
Transcriptome-based characterization of the interactions between Saccharomyces cerevisiae and Lactobacillus delbrueckii subsp. bulgaricus in lactose-grown chemostat co-cultures |
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