|
| Status |
Public on Jun 05, 2018 |
| Title |
CAMHB + 0.5 ug/mL Nafcillin Rep 3 |
| Sample type |
SRA |
| |
|
| Source name |
Staphylococcus aureus
|
| Organism |
Staphylococcus aureus |
| Characteristics |
strain: USA300 TCH1516
media: CAMHB
nafcillin concentration (ug/ml): 0.5
|
| Growth protocol |
Overnight cultures of CA-MRSA TCH1516 were diluted to a starting OD600 of 0.01 into fresh media and grown at 37C with stirring to OD600 of 0.4. This preculture was then diluted back to OD600 0.01 into fresh media containing no drug or sub-inhibitory concentrations of nafcillin relative for each media type. Growth was monitored by obtaining OD600 readings every 30 min and sample for RNA sequencing was taken at 2.5 hours.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
After 2.5 hours of growth, 3 mL samples were taken for RNA sequencing and added to tubes containing 6 mL RNAprotect. After incubation, they were centrifuged to remove the supernatant. RNA was extracted from the pelleted cells using a ‘Quick RNA Fungal/Bacterial Microprep’ kit developed by Zymo Research. The cells were mechanically lysed with a Roche MagNa Lyzer instrument and DNA was removed with DNase I during the RNA purification. RNA quality was checked with an Agilent Bioanalyzer instrument and ribosomal RNA was removed using an Illumina Ribo-Zero kit.
The remaining RNA was used to build a cDNA library for sequencing using a KAPA Stranded RNA-seq Library Preparation Kit. The kit was used for RNA fragmentation, sequencing adapter ligation, and library amplification. The generated cDNA libraries were sent for Illumina sequencing on a HiSeq 4000.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 4000 |
| |
|
| Data processing |
Sequenced reads were aligned to TCH1516 genome (NC_010079.1) using bowtie-2 version 2.2.6 with parameters ’ -X 1000 -N 1 -3 3 ‘
Gene specific counts were generated from aligned reads with HTSeq version 0.9.1 using parameters ‘-s reverse -r pos’
Normalized TPM counts were generated with DESEq2 1.16.1
ComBat module in SVA package version 3.24.4 was used for batch correction of TPM data
Genome_build: NC_010079.1
Supplementary_files_format_and_content: Tab delimited file with read counts, normalized TPM counts and batch corrected TPM counts
|
| |
|
| Submission date |
Jun 04, 2018 |
| Last update date |
Jun 05, 2018 |
| Contact name |
Saugat Poudel |
| E-mail(s) |
sapoudel@ucsd.edu
|
| Organization name |
University of California San Diego
|
| Street address |
9500 Gilman Dr.
|
| City |
La Jolla |
| ZIP/Postal code |
92037 |
| Country |
USA |
| |
|
| Platform ID |
GPL25144 |
| Series (1) |
| GSE115309 |
Staphylococcus aureus TCH1516 response to sub-inhibitory concentrations of Nafcillin in different media conditions |
|
| Relations |
| BioSample |
SAMN08200716 |
| SRA |
SRX4112977 |
