|
| Status |
Public on Aug 28, 2017 |
| Title |
LT2 ridA Rep 2 |
| Sample type |
SRA |
| |
|
| Source name |
cell lysate
|
| Organism |
Salmonella enterica subsp. enterica serovar Typhimurium str. LT2 |
| Characteristics |
strain: DM3480
genotype: ridA
|
| Treatment protocol |
Growth in minimal medium was the only treatment condition
|
| Growth protocol |
S. enterica LT2 WT and ridA strains were subcultured from overnight cultures grown in nutrient broth medium and subcultured 1:100 in 5 mL no-carbon E medium supplemented with 1 mM MgSO4, trace minerals, and 11mM D-glucose and grown at 37C shaking until to mid-log (OD650 = 0.6)
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Each 5 mL sample was quickly centrifuged at 16,000 x g in 1.5 mL Eppendorf tubes, supernatant removed and pellets flash-frozen in liquid nitrogen and kept on dry ice. Total RNA was extracted using the RNAsnapTM method (Stead, et al., 2012). RNA was subjected to an ethanol precipitation and DNA/RNA pellets were resuspended in 90 μl UltraPureTM water at 4°C overnight prior to treatment with RNase-free Turbo DNase (Ambion), and being precipitated once more by sodium acetate-ethanol treatment. Resulting RNA was resuspended at 4°C for 3 h and stored at -80°C until use. The total RNA was extracted from four independent replicates of S. enterica wild-type and ridA mutant strains. In each of the resulting eight samples, rRNA was removed from 5 μg total RNA using the Ribo-Zero rRNA removal kit (Bacteria; Illumina) following manufacturer’s recommendations.
A cDNA library was created from samples containing <10% rRNA contamination, as measured by the RNA 6000 pico kit for the Agilent 2100 bioanalyzer, and sequenced by the University of Georgia Genomics Facility (GGF) from these RNA samples. Specifically, one hundred nanograms of the rRNA-free RNA was used to create each sequencing library using the KAPA stranded RNA-seq kit (KAPA Biosystems).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
rRNA was removed from 5 μg total RNA using the Ribo-Zero rRNA removal kit (Bacteria, Illumina)
|
| Data processing |
Real-Time Analysis v2 software used for basecalling.
Data Processing steps were performed by Walter Lorenz of the Quantitative Biology Consulting Group at the University of Georgia after data was downloaded from the Georgia Genomics Facility BaseSpace link.
Sequenced reads were trimmed by Trimmomatic for adaptor sequence, and masked for low-complexity or low-quality sequence, and only trimmed data that was maintained as paired data was used to map to the S. enterica LT2 genome using EdgePro version 1.3.1
The matrix files generated by EdgePro were imported into Deseq in order to perform statistical analysis for determination of differentially expressed genes (FDR > 0.05).
Genome_build: ASM694v1
Supplementary_files_format_and_content: SE_Deseq_Processed.xlsx is an excel file format file generated by the Quantitative Biology Consulting Group at the University of Georgia and provided to the researchers in this study. It contains raw and normalized reads generated by Deseq as well as FDR values for all LT2 ORFs given as well as a calculated fold-change number for each gene between the wild-type and ridA strains
|
| |
|
| Submission date |
Aug 27, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Diana M Downs |
| E-mail(s) |
dmdowns@uga.edu
|
| Organization name |
University of Georgia
|
| Department |
Microbiology
|
| Lab |
Downs
|
| Street address |
527 Biological Sciences Building
|
| City |
Athens |
| State/province |
GA |
| ZIP/Postal code |
30602 |
| Country |
USA |
| |
|
| Platform ID |
GPL23956 |
| Series (1) |
| GSE103146 |
Determination of S. enterica wild-type and ridA mutant Transcriptome Differential Expression |
|
| Relations |
| BioSample |
SAMN07563781 |
| SRA |
SRX3136380 |
