|
| Status |
Public on Dec 11, 2020 |
| Title |
A498_E. coli 83972_exp1 |
| Sample type |
RNA |
| |
|
| Source name |
A498 kidney cells treated with E. coli 83972 supernatant
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: A498
treatment: E. coli 83972 supernatant
concentration: 10^8 CFU/ml
|
| Treatment protocol |
Cell culture medium was aspirated and replaced with fresh serum-free RPMI. E. coli strains were diluted in PBS and added to the cells at a concentration of 10^8 CFU/ml for 4 hours. Alternatively, cells were treated with bacterial supernatant or treated with recombinant protein (50, 100 or 250 ug/ml) for 4 hours. The control sample was treated with a similar volume of PBS.
|
| Growth protocol |
E. coli strains were grown on tryptic soy agar (16 h, 37˚C). The human epithelial cell line A498 (ATCC HTB44, human kidney carcinoma) was cultured in RPMI-1640 supplemented with 1 mM sodium pyruvate, 1 mM non-essential amino acids and 10% fetal calf serum. Bacteria activation of the cells was studied on confluent cell layers in 6-well tissue culture plates (Nunc). A498 cells on confluent cell layers in 6-well tissue culture plates were treated. For bacterial supernatant, 10^8 CFU/ml of bacteria was grown in serum-free RPMI for 4 hours.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using the RNeasy Mini Kit (Qiagen) and on-column DNase digestion, according to the manufacturer's instructions.
|
| Label |
biotin
|
| Label protocol |
100 ng of total RNA was amplified, labeled and fragmented using the GeneChip 3´IVT PLUS Kit (ThermoFisher Scientific). Biotinylated cRNA was prepared according to the standard Affymetrix's protocol.
|
| |
|
| Hybridization protocol |
6ug of fragmented and labelled aRNA was hybridized onto Human Genome U219 array strips (Affymetrix) for 16 hours at 45 °C, washed, stained (Applied Biosystems, ThermoFisher Scientific).
|
| Scan protocol |
Scanning was done using the GeneAtlas system (Affymetrix).
|
| Description |
Gene expression data from 83972 supernatant treated A498 cells
|
| Data processing |
Transcriptomic data was normalized using Robust Multi Average implemented in the Transcriptome Analysis Console software (v.4.0.1.36, Applied Biosystems, ThermoFisher Scientific).
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| |
|
| Submission date |
Nov 25, 2020 |
| Last update date |
Dec 12, 2020 |
| Contact name |
Ines Ambite |
| E-mail(s) |
[email protected]
|
| Organization name |
Lund University
|
| Department |
Laboratory Medicine
|
| Lab |
MIG
|
| Street address |
Klinikgatan 28
|
| City |
Lund |
| ZIP/Postal code |
22184 |
| Country |
Sweden |
| |
|
| Platform ID |
GPL13667 |
| Series (1) |
| GSE162190 |
Expression data profile of A498 cells infected with E. coli 83972, mutants or stimualted with rNlpD |
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