|
| Status |
Public on Jun 19, 2017 |
| Title |
sulfide-treated [HS2] |
| Sample type |
SRA |
| |
|
| Source name |
Exponentially grown cells
|
| Organism |
Staphylococcus aureus |
| Characteristics |
strain: Newman
genotype/variation: WT
treatment: sodium sulfide
|
| Treatment protocol |
Na2S was added to the cultures at a final concentration of 0.2 mM when the OD600 reached 0.2.
|
| Growth protocol |
Overnight starter cultures of S. aureus strain Newman were grown in TSB medium overnight and used to inoculate 15mL cultures in Hussain−Hastings−White modified (HHWm) minimal media (Toledo-Arana et al., 2005) supplemented with 0.5 mM thiosulfate and 0.5 mM methionine at a starting OD600 = 0.02. All cultures were grown in loosely capped 50mL Falcon tubes at 37°C with shaking at 200 rpm, until a final OD600 of 0.2.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Following growth to OD600=0.2, 5mL aliquots of cells were pelleted and resuspended in 1mL TriReagent (Molecular Research Center), then lysed with 0.1mm silica beads in a bead beater (MP Bio). RNA was extracted by adding 200 uL chloroform, mixing, and centrifuging. The aqueous layer was extracted and added to one volume of 70% ethanol. RNA purification was completeted using the RNeasy minikit (Qiagen).
A total of 1 μg of RNA was used to make stranded, rRNA-delpeted RNAseq libraries using the ScriptSeq Complete (Bacteria) kit (then from Epicentre, now Illumnia Cat. # BB1206).
Sequencing was performed using an Illumina HiSeq2000 instrument with a read length of 1 × 50 nucleotides. Image analysis was done by the Illumina HCS.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
countstable.txt.gz
HS2
|
| Data processing |
Bases were called using the Illumina HCS
Sequencing reads were trimmed using Trimmomatic (version 0.33; 1) with the following parameters: ILLUMINACLIP:adapter.fa:2:20:6 LEADING:3 TRAILING:3 SLIDINGWINDOW:4:20 MINLEN:35.
The trimmed reads were mapped on to the Staphylococcus aureus subsp. aureus str. Newman genome using bowtie2 (version 2.1 with default parameters; 2).
Read counts for genes and intergenic intervals were calculated using a custom perl script. Resulting gene/interval counts were used to conduct differential expression analysis using the program DESeq2 algorithm (3) with default parameters.
Calprotectin data quality was checked with FastQC and QC3. Alignment was performed by TopHat. Gene quantification (FPKM) was done using Cufflinks. Differential expression analysis was carried out using Cuffdiff from the Cufflinks package.
Genome_build: AP009351.1
Supplementary_files_format_and_content: raw counts of sequencing reads for the features of interest including genes and intergenic regions for each fastq file; intergenic regions are identified as follows Inter.<molecule>.<strand>.<basepair_begin>.<basepair_end>
|
| |
|
| Submission date |
May 30, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
David Peter Giedroc |
| Organization name |
Indiana University
|
| Department |
Chemistry
|
| Lab |
Simon Hall 320C
|
| Street address |
212 S. Hawthorne Drive
|
| City |
Bloomington |
| State/province |
IN |
| ZIP/Postal code |
47405 |
| Country |
USA |
| |
|
| Platform ID |
GPL17452 |
| Series (1) |
| GSE99432 |
Changes in relative transcript amounts caused by hydrogen sulfide treatment, calprotectin treatment, and deletion of CstR in Staphylococcus Aureus |
|
| Relations |
| BioSample |
SAMN07177374 |
| SRA |
SRX2868736 |
