|
| Status |
Public on Sep 11, 2020 |
| Title |
Cell aggregates of S. Typhimurium UMR1 treated with 15 µM clarithromycin [4_S4] |
| Sample type |
SRA |
| |
|
| Source name |
Rdar Biofilm-Forming Bacteria
|
| Organism |
Salmonella enterica subsp. enterica serovar Typhimurium |
| Characteristics |
strain: ATCC14028 Nalr (strain UMR1)
treatment: 15 µM clarithromycin
|
| Growth protocol |
Prior to the experiment, single colonies of bacteria were inoculated in 5 mL LB and grown overnight at 37 ˚C with shaking. Subsequently, strains were inoculated to OD600 0.02 and grown in Luria broth (LB) without salt medium under microaerophilic conditions (flask with 60% medium filling level) with shaking at 150 rpm at 28 ºC for 16 h.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Cells were harvested by centrifugation and disrupted by lysis buffer (50 mM Tris HCL pH 8, 8 % sucrose, 0.5 % Triton, 20 mM EDTA, 4 mg/mL), 200 µL acidic phenol was added, and samples were then heated at 65 ºC for 15 min, while shaking. The sample was cooled on ice for 5 min and centrifuged at 13000 rpm. The aqueous phase was collected and extracted once with an acid phenol:chloroform mix, and twice with chloroform. Finally, RNA was precipitated with 3 volumes of 100 % ethanol and 0.3 M sodium acetate overnight at -80 ºC. Pellets were washed with 1 mL 70 % ethanol, and air-dried after the supernatant was removed. Pellet was dissolved in 100 µL DEPC-treated distilled water. After DNase (Ambion RiboPure-Yeast DNase) treatment, the RNA samples were analysed by denaturating gel electrophoresis to assess DNA contamination and integrity of the isolated RNA. If DNA contamination was still present, the RNA was repeatedly incubated with DNAse. The RNA concentration was measured by Nano drop and the quality of RNA preparation assessed by Agilent Bioanalyzer 2100. The DNA-free RNA was stored at -80 ºC until use.
RNA samples were sequenced on a Nextseq 550 with medium output flowcell 150 cycles (2x75 PE) after depletion from ribosomal RNAs by the RiboMinus Transcriptome Isolation Kit (Invitrogen).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
NextSeq 550 |
| |
|
| Description |
4_S4
Treated
|
| Data processing |
Bcl files produced by NexSeq 550 were converted to fastq and demultiplexed using the bcl2fastq v2.20.0.422 program
Fastq files were aligned to the Salmonella enterica subsp. enterica serovar Typhimurium str. 14028S (NC_016856.1) genome using STAR v2.5.2b
Reads were then counted in annotated genes using featureCounts v1.5.1
Genome_build: NC_016856.1
Supplementary_files_format_and_content: counted reads in annotated genes
|
| |
|
| Submission date |
Sep 09, 2020 |
| Last update date |
Sep 11, 2020 |
| Contact name |
Ute Römling |
| E-mail(s) |
ute.romling@ki.se
|
| Organization name |
Karolinska Institutet
|
| Department |
Department of Microbiology, Tumor and Cell Biology
|
| Street address |
Solnavägen 9
|
| City |
Stockholm |
| ZIP/Postal code |
17177 |
| Country |
Sweden |
| |
|
| Platform ID |
GPL28222 |
| Series (1) |
| GSE157695 |
An antibiofilm effect against Salmonella typhimurium rdar biofilm formation upon treatment with the macrolide clarithromycin |
|
| Relations |
| BioSample |
SAMN16086420 |
| SRA |
SRX9098773 |
