|
| Status |
Public on Dec 22, 2020 |
| Title |
USA400 MW2 Biofilm 10 hour |
| Sample type |
SRA |
| |
|
| Source name |
bacterial cells
|
| Organism |
Staphylococcus aureus |
| Characteristics |
strain: MW2
genotype: Wildtype
growth medium: TSB
growth conditions: Static biofilm, 10 h, 37°C
|
| Treatment protocol |
Samples were stored at -80°C prior to RNA extraction.
|
| Growth protocol |
Overnight S. aureus cultures were used to inoculate fresh tryptic soy broth at an optical density (OD600) of 0.5 in 96-well microtiter plates, in biological triplicate. Test cultures were grown for 5 h, 10 h and 24 h at 37°C in a static incubator prior to collection. To collect planktonic samples, 75 μL of supernatant was removed from the top of each well, and those for like strains were pooled. Samples were immediately combined with 5 mL of ice-cold PBS, and pelleted by refrigerated centrifugation for subsequent RNA isolation. For biofilm samples, the remaining supernatant was removed and biofilm containing wells were washed three times with 200 μL of ice-cold PBS. Ice-cold PBS was added a final time, pipetted vigorously to disrupt biofilm cells, and like strains were pooled. Samples were then immediately combined with an additional 5 mL of ice-cold PBS and pelleted by refrigerated centrifugation for subsequent RNA isolation.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total bacterial RNA was isolated utilizing an RNeasy Kit (Qiagen) and contaminating DNA was removed by treatment with TURBO DNA-free kit (Ambion). Samples with a RIN of >9.7 measured by Agilent 2100 Bioanalyzer system and corresponding RNA 6000 Nano kit (Agilent) were used in this study. Ribosomal RNA was removed using a Ribo-Zero Kit for Gram-positive Bacteria (Illumina) and MICROBExpress Bacterial mRNA enrichment kit (Agilent).
Library construction was performed using the TruSeq Stranded mRNA Kit from Illumina omitting mRNA enrichment steps.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
Raw data files (.fastq) were exported from the Illumina BaseSpace Server
Raw data files (.fastq) were imported into Qiagen CLC Genomics Workbench (version 20) as paired-end reads for analysis and adapter indices trimmed.
Reads corresponding to ribosomal RNA were filtered out and removed
Gene expression values were calculated using the Expression Browser tool
Differential expression between biofilm and planktonic samples were generated for each strain and timepoint individually using the Differential Expression in Two Groups tool
Fold change comparisons were exported in excel format
Genome_build: The accession number for the Genbank genome files used as reference are: USA100 (NC_002745), USA200 (NC_002952), USA300 (NC_007793), USA400 (NC_003923), and USA500 (NC_007793).
Supplementary_files_format_and_content: Differential expression in biofilm populations relative to their planktonic counterpart are presented as fold-change values in excel format.
|
| |
|
| Submission date |
Dec 14, 2020 |
| Last update date |
Dec 22, 2020 |
| Contact name |
Lindsey N Shaw |
| Organization name |
University of South Florida
|
| Department |
CMMB
|
| Street address |
4202 E Fowler Avenue
|
| City |
Tampa |
| State/province |
FL |
| ZIP/Postal code |
33620 |
| Country |
USA |
| |
|
| Platform ID |
GPL24034 |
| Series (1) |
| GSE163153 |
Temporal transcriptomic analysis of five Staphylococcus aureus strains' biofilms and planktonic counterparts |
|
| Relations |
| BioSample |
SAMN17077274 |
| SRA |
SRX9682660 |
