|
| Status |
Public on Sep 22, 2008 |
| Title |
multi-bicarb-v-ko-bicarb-1 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
multi-bicarb
|
| Organism |
Citrobacter rodentium |
| Characteristics |
RegA multicopy transgene
agent: bicarbonate
|
| Treatment protocol |
Bicarbonate was added to a final concentration of 45mM
|
| Growth protocol |
overnight cultures of Citrobacter rodentium were inoculated into LB medium to obtain an OD600 of 0.1 and grown until the OD600 was 0.88
|
| Extracted molecule |
total RNA |
| Extraction protocol |
10mL of a culture was incubated with 20mL of RNAprotect solution (Qiagen) at room temperature for 15min. Cells were pelleted and RNA was purified using a FastRNA Pro Blue kit (Qbiogene). RNA samples were Dnase treated (Qiagen) and further purified using an RNeasy MiniElute kit (Qiagen).
|
| Label |
Cy3
|
| Label protocol |
5microg of total RNA was labeled with Cy3-ULS or Cy5-ULS (Kreatech) as per manufacturers instructions
|
| |
|
| Channel 2 |
| Source name |
ko-bicarb
|
| Organism |
Citrobacter rodentium |
| Characteristics |
agent: bicarbonate
RegA Knockout
|
| Treatment protocol |
Bicarbonate was added to a final concentration of 45mM
|
| Growth protocol |
overnight cultures of Citrobacter rodentium were inoculated into LB medium to obtain an OD600 of 0.1 and grown until the OD600 was 0.88
|
| Extracted molecule |
total RNA |
| Extraction protocol |
10mL of a culture was incubated with 20mL of RNAprotect solution (Qiagen) at room temperature for 15min. Cells were pelleted and RNA was purified using a FastRNA Pro Blue kit (Qbiogene). RNA samples were Dnase treated (Qiagen) and further purified using an RNeasy MiniElute kit (Qiagen).
|
| Label |
Cy5
|
| Label protocol |
5microg of total RNA was labeled with Cy3-ULS or Cy5-ULS (Kreatech) as per manufacturers instructions
|
| |
|
| |
| Hybridization protocol |
Hybridization was conducted as per Agilent 2 color microarray-based gene expression analysis manual (version 5.7) with the modification 4 microg of labeled total RNA was used per hybridization.
|
| Scan protocol |
Images were quantified using Agilent Feature Extraction Software.
|
| Description |
no additional information
|
| Data processing |
Data was normalized within arrays using loess normalization in limma. DE was analyzed in limma with design <- cbind(dye=c(1,1,1,1,1,1,1,1), UnstimKOvMC=c(0,-1,0,-1,0,1,0,1), StimKOvMC=c(-1,0,-1,0,1,0,1,0))
VALUE column below reports normalized log2 (ko/ref) ratios.
|
| |
|
| Submission date |
Sep 22, 2008 |
| Last update date |
Sep 22, 2008 |
| Contact name |
Matthew J Wakefield |
| E-mail(s) |
[email protected]
|
| Organization name |
Walter and Eliza Hall Institute
|
| Department |
Bioinformatics
|
| Street address |
1G Royal Parade
|
| City |
Parkville |
| State/province |
VIC |
| ZIP/Postal code |
3050 |
| Country |
Australia |
| |
|
| Platform ID |
GPL7345 |
| Series (1) |
| GSE12876 |
Control of Virulence Gene Expression in Citrobacter rodentium by Trascription Factor RegA |
|
