|
| Status |
Public on Dec 01, 2015 |
| Title |
Peg(-0.5)_1 |
| Sample type |
RNA |
| |
|
| Source name |
Matric stress -0.5Mpa
|
| Organism |
Pseudomonas veronii 1YdBTEX2 |
| Characteristics |
media: 21C minimal medium with 5mM succinate and PEG8000 at -0.5Mpa
|
| Treatment protocol |
10 ml of culture sample was added to 10 ml of 21CS medium containing PEG8000 so as to obtain a final matric potential of -0.5 Mpa or 10ml of 21CS medium containing NaCl to obtain a final solute potential of -0.5Mpa and incubated for 30 min. Each treatment or control was run in independent replicates.
|
| Growth protocol |
Pseudomonas veronii (1YdBTEX2) was cultured in 21C medium with succinate 5mM as carbon source at 30°C and 180 rpm until exponential phase (OD600= 0.3)
|
| Extracted molecule |
total RNA |
| Extraction protocol |
After 30 min incubation samples were immediately filtered over a 0.2 µm membrane filter by vacuum suction. The filter with the cells was removed and frozen in liquid nitrogen. The filters were stored at -80°C until RNA isolation. RNA was extracted from frozen cells on filters with an acid-phenol method.
|
| Label |
Cyanine 3-dCTP
|
| Label protocol |
Labelled cDNAs were produced in a reverse transcription reaction using cyanine-3-labeled dCTP. The labelled cDNA was purified with a MinElute PCR purification kit (Qiagen 28004). The percentage of incorporation of Cy-3-dCTP was calculated by the MICROARRAY function of the Nanodrop spectrophotometer.
|
| |
|
| Hybridization protocol |
The volume of the different samples was adjusted for the hybridization in order to add 60 ng of labelled Cy3-cDNA per array on the slide. Hybridization was performed at 65°C for 17 hours, after which slides were washed according to Agilent procedures and scanned. The fragmentation step (heating to 60°C for 30 minutes) was omitted.
|
| Scan protocol |
Microarray slides were scanned using an Agilent High-Resolution Microarray Scanner (G2505), immediately after hybridization using the One-Color microarray -based gene expression analysis protocol provided by Agilent (scan area 61x21.6mm XDRHi:100% XDRLo:10%). The AGILENT FEATURE EXTRACTION SOFTWARE was used to extract the signal intensities of the probes from the scanned images.
|
| Description |
Hot-phenol extraccion
|
| Data processing |
The text data file was used as input in GeneSpring GX v12. Expression data were quantile normalized by GeneSpring and baseline transformed. All genes were filtered by expression and were retained when the signal intensity was above the 20th percentile in at least one of the samples.
|
| |
|
| Submission date |
May 19, 2015 |
| Last update date |
Dec 01, 2015 |
| Contact name |
Silvia K. Moreno-Forero |
| E-mail(s) |
[email protected]
|
| Organization name |
University of Lausanne
|
| Department |
Department of Fundamental Microbiology
|
| Street address |
Batiment Biophore, Unil Sorge
|
| City |
Lausanne |
| State/province |
Vaud |
| ZIP/Postal code |
1015 |
| Country |
Switzerland |
| |
|
| Platform ID |
GPL20216 |
| Series (1) |
| GSE69022 |
Transcriptional responses of Pseudomonas veronii 1YdBTEX2 to the exposure to laboratory induced matric and solute stresses |
|
