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| Status |
Public on Jul 31, 2024 |
| Title |
bacteria, OD1, subculture, adsa-Proline R2 |
| Sample type |
SRA |
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| Source name |
Newman
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| Organism |
Staphylococcus aureus |
| Characteristics |
tissue: Newman
cell line: NC_009641.5
genotype: delta-adsA
treatment: luria broth + proline
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| Treatment protocol |
treatment of cohort B and D in 100uM proline
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| Growth protocol |
overnight in luria borth or luria broth + 100uM proline, subcultured and grown to OD1 ebfore extraction
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| Extracted molecule |
total RNA |
| Extraction protocol |
Bacterial pellets were incubated in a cell wall lysis mixture TE buffer pH 8 (30 mMTris, 1mMEDTA, 15 mg/mL and 200μg/mL proteinase K) containing mutanolysin, lysostaphin and lysozyme at 37 °C for 30 minutes. TRK lysis buffer and 70% ethanol were added to each sample, prior to transferring to E.Z.N.A RNA isolation columns. RNA was isolated following the manufacturer’s instructions and DNA was selectively degraded using the DNA-free DNA removal kit.The RNA was precipitated with 0.1 volume 3 M sodium acetate (ThermoFisher #S209) and 3 volumes of 100% ethanol, recovered by centrifugation and washed with ice cold 70% ethanol. A ribosomal RNA-depleted cDNA library was prepared according to the manufacturer’s instructions using the Universal Prokaryotic RNA-Seq Prokaryotic AnyDeplete kit (NuGEN #0363-32) and sequenced with Illumina HiSeq.
Raw base calls were converted to fatsq files using Bcl2fastqs. Filtered reads were aligned to the Newman (Refseq NC_009641.1) reference genome using STAR-Aligner v2.7.3a. The mapped reads were annotated for read groups and marked for duplicates using the Picard tools v2.22.3.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 3000 |
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| Data processing |
The raw counts were quantified using Subreads:FeatureCounts v1.6.3 and processed for differential gene expression using DEseq2 in R v3.5.3.
Analysis of sequencing for clinical isolates was performed by Windmüller and colleagues. Briefly, Data was stratified into ≥2f, ≥4f, ≥8f and into genes that were either not expressed in both isolates or not present on the genome level in both isolates. To exclude low abundant transcripts, a threshold of less than 3 reads per gene was used.
Assembly: NC_009641.1
Supplementary files format and content: processed data in txt files and csv
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| Submission date |
May 29, 2024 |
| Last update date |
Jul 31, 2024 |
| Contact name |
ALICE PRINCE |
| E-mail(s) |
ASP7@cumc.columbia.edu
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| Organization name |
COLUMBIA UNIVERSITY
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| Department |
PEDIATRICS ID
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| Lab |
PRINCE LAB
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| Street address |
650 W 168TH ST
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| City |
NEW YORK |
| State/province |
NY |
| ZIP/Postal code |
10032 |
| Country |
USA |
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| Platform ID |
GPL26273 |
| Series (1) |
| GSE268637 |
Staphylococcus aureus adapts to exploit collagen-derived proline during chronic infection |
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| Relations |
| BioSample |
SAMN41592601 |
| SRA |
SRX24744539 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM8295812_ADSAP2.counts.txt.gz |
1.2 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
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