|
| Status |
Public on Sep 11, 2014 |
| Title |
WT_noCPR_1 |
| Sample type |
RNA |
| |
|
| Source name |
Non-treated, WT control at T0, biological rep 1
|
| Organism |
Escherichia coli K-12 |
| Characteristics |
background strain: BW25113
genotype/variation: wild type
|
| Treatment protocol |
We compared the microarray-determined mRNA profiles (Affymetrix E. coli Genome 2.0 Array) of Δfur and WT genotypes in response to ciprofloxacin treatment (100 ng/ml) with those of untreated cultures. We used two biological replicates per genotype.
|
| Growth protocol |
Escherichia coli BW25113 and fur mutant strains were cultivated at 37Co in 25 ml of LB medium using Erlenmeyer flasks. 1 ml of OVN culture was transferred to 20 ml of fresh LB medium and cells were incubated in the presence or absence of ciprofloxacin (100 ng/ml).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Samples for microarray analyses were taken before ciprofloxacin treatment (t0) and then at 1 hour post-treatment (t1). Cell concentration was adjusted to 109 cells/ml and cultures were treated with ciprofloxacin in Erlenmeyer flasks using 20 ml fresh LB. ~109 cells were taken at each timepoint as samples for RNA isolation. In order to stabilize RNA, RNA Bacteria Protect Reagent (Qiagen) was added to the samples according to the instructions of the manufacturer and then samples were stored overnight at -80°C. Total RNA was isolated by using RNeasy Mini Kit (Qiagen) according to the manufacturer’s instructions. Sample concentration was estimated using the NanoDrop 1000 (Thermoscientific) spectrophotometer.
|
| Label |
biotin
|
| Label protocol |
Biotinylated cDNA were prepared according to the standard Affymetrix protocol from 6 ug total RNA (Expression Analysis Technical Manual, 2001, Affymetrix).
|
| |
|
| Hybridization protocol |
Hybridization of the cDNAs was conducted at UD-GenoMed Medical Genomic Technologies Ltd. (4032, Debrecen, Nagyerdei Krt. 98. Tel: +36 30 494 6496) using Affymetrix Genechip® E. coli Genome 2.0 Array (Affymetrix) as described by the manufacturer.
|
| Scan protocol |
Arrays were scanned on GeneChip Scanner 7G (Affymetrix), Affymetrix GeneChip Command Console (AGCC) was used to obtain image files (DAT/CEL).
|
| Description |
Escherichia coli liquid culture, incubated in LB medium, BW25113
|
| Data processing |
CHP files were generated using Affymetrix Expression Control. Downstream data analysis was performed in GeneSpring GX 12, CEL files were imported using RMA algorithm and median normalization was performed.
|
| |
|
| Submission date |
Mar 06, 2014 |
| Last update date |
Sep 11, 2014 |
| Contact name |
Balazs Bogos |
| E-mail(s) |
bogos.balazs@mta.brc.hu
|
| Phone |
+36-62-599-693
|
| Organization name |
Biological Research Centre of the Hungarian Academy of Sciences
|
| Department |
Institute of Biochemistry
|
| Lab |
Laboratory of Microbial Experimental Evolution
|
| Street address |
Temesvári krt. 62-64
|
| City |
Szeged |
| ZIP/Postal code |
6724 |
| Country |
Hungary |
| |
|
| Platform ID |
GPL3154 |
| Series (1) |
| GSE55662 |
Plasticity of transcriptional regulation under antibiotic stress |
|
