|
| Status |
Public on Aug 20, 2016 |
| Title |
Strain BP536 Mg modulated versus BP536 replicate 1 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
pellet of bacteria
|
| Organism |
Bordetella pertussis |
| Characteristics |
strain: BP536
genotype: Wild-type
modulation mg: No
|
| Growth protocol |
Bordetella pertussis strains were first grown on Bordet-Gengou (BG) agar plates and in Stainer-Scholte (SS) liquid medium supplemented with 100 µg/ml streptomycin
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA extracted using TriReagent (Ambion) following manufacturer's instructions
|
| Label |
Cy3
|
| Label protocol |
15 µg of total RNA was reverse transcribed with 400 units of SuperScript III (Invitrogen) in presence of 100µM Cy3-dCTP or Cy5-dCTP (GE) and 300 mM of hexanucleotide random (Roche) during 2 hours à 42°C. The labelled cDNA was then NaOH treated to degrade RNA and purified on Qiaquick PCR purification kit (Qiagen) according to the manufacturer’s instructions.
|
| |
|
| Channel 2 |
| Source name |
pellet of bacteria
|
| Organism |
Bordetella pertussis |
| Characteristics |
strain: BP536
genotype: Wild-type
modulation: Yes
|
| Growth protocol |
Bordetella pertussis strains were first grown on Bordet-Gengou (BG) agar plates and in Stainer-Scholte (SS) liquid medium supplemented with 100 µg/ml streptomycin
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA extracted using TriReagent (Ambion) following manufacturer's instructions
|
| Label |
Cy5
|
| Label protocol |
15 µg of total RNA was reverse transcribed with 400 units of SuperScript III (Invitrogen) in presence of 100µM Cy3-dCTP or Cy5-dCTP (GE) and 300 mM of hexanucleotide random (Roche) during 2 hours à 42°C. The labelled cDNA was then NaOH treated to degrade RNA and purified on Qiaquick PCR purification kit (Qiagen) according to the manufacturer’s instructions.
|
| |
|
| |
| Hybridization protocol |
Hybridization was performed in 40% formamide, 5x Denhardt’s solution, 0.1% SDS, 1 mM Sodium pyrophosphate and 5x SSC during 14-16h at 52°C under agitation in rotating oven. Slides were then washed sequentially in 2× SSC/0.2% SDS during 5 min, 0.5× SSC during 10 min, 0.05x SSC during 5 min and 0.01x SSC during 1 min at room temperature before drying.
|
| Scan protocol |
Scanned on an Innoscan 700 scanner (Innopsys), images were segmented using Mapix Software (version 3.2.1).
|
| Description |
BP536 vs BP536 Mg comparison
|
| Data processing |
For normalization and differential expression analyses the LIMMA package (Linear Models for Microarray Data), running under the statistical language R v2.15.0, was used. Statistically significant regulation was identified using moderated t-statistic with empirical Bayes shrinkage of the standard errors. Because of multiple testing, obtained P-values were corrected using Benjamini & Hochberg method to control false discovery rates.
|
| |
|
| Submission date |
Feb 09, 2016 |
| Last update date |
Aug 20, 2016 |
| Contact name |
David Hot |
| E-mail(s) |
[email protected]
|
| Organization name |
Institut Pasteur de Lille
|
| Department |
Center for Infection and Immunity of Lille
|
| Lab |
Transcriptomics and Applied Genomics
|
| Street address |
1 rue du professeur Calmette
|
| City |
Lille |
| ZIP/Postal code |
59000 |
| Country |
France |
| |
|
| Platform ID |
GPL19266 |
| Series (1) |
| GSE77754 |
The multifaceted RisA regulon of Bordetella pertussis |
|
