|
| Status |
Public on Sep 01, 2022 |
| Title |
WalK T389A mutant replicate 2 |
| Sample type |
SRA |
| |
|
| Source name |
USA300
|
| Organism |
Staphylococcus aureus |
| Characteristics |
time: mid-exponential
culture medium: TSB broth
growth temp: 37C
strain: USA300
genotype: WalK T389A
|
| Treatment protocol |
A 5 ml aliquot of culture was removed and added to 10 ml of RNAprotect Bacteria Reagent (Qiagen) and mixed by vortexing. The sample was incubated at room temperature for 5 min. Cells were then harvested by centrifugation (7,000xg/5min/22°C), the supernatant discarded and the cell pellet resuspended in 1 ml of TRIzol (Invitrogen).
|
| Growth protocol |
A 10 ml TSB (50 ml tube) culture was grown overnight at 37°C with shaking at 200rpm. The saturated culture was diluted 1:100 into fresh 10 ml TSB and grown to an OD600nm of 0.8-1.0
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Cells were lysed by bead beating (Precellys 24 instrument - 6000 rpm, 1 min, 100 um zirconium beads) and cell lysates were clarified by centrifugation (20,000xg/10 min/4°C). Subsequently, 700 µl of supernatant was removed and mixed with 700 µl of ethanol. RNA was extracted using a Direct-Zol RNA miniprep plus kit (ZymoResearch) according to manufactures instructions, including the on column DNase I treatment step.
The libraries were sequenced on the Illumina HiSeq platform for 50 bp single end reads.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Data processing |
RNAseq data was analysed using the S. aureus USA300 FPR3757 reference genome (accession number: NC_007793) and Kallisto, a kmer based pseudoalignment tool, with analysis and visualization using Degust [https://github.com/drpowell/degust]
Assembly: S. aureus USA300 NC_007793
|
| |
|
| Submission date |
Aug 30, 2022 |
| Last update date |
Sep 01, 2022 |
| Contact name |
Tim Stinear |
| E-mail(s) |
tstinear@unimelb.edu.au
|
| Organization name |
University of Melbourne
|
| Street address |
792 Elizabeth St, Melbourne
|
| City |
Melbourne |
| State/province |
Victoria |
| ZIP/Postal code |
3000 |
| Country |
Australia |
| |
|
| Platform ID |
GPL17452 |
| Series (2) |
| GSE212319 |
High resolution definition of the Staphylococcus aureus WalKR regulon [RNA-seq] |
| GSE212321 |
High resolution definition of the Staphylococcus aureus WalKR regulon |
|
| Relations |
| BioSample |
SAMN30594101 |
| SRA |
SRX17321358 |
