|
| Status |
Public on Oct 25, 2021 |
| Title |
mexR*-Exponential |
| Sample type |
SRA |
| |
|
| Source name |
Exponential growth phase in LB liquid cultures
|
| Organism |
Pseudomonas aeruginosa |
| Characteristics |
strain: JFL30 (mexR*)
genotype/variation: MexAB-OprM overexpressing mutant
type of culture: Planktonic
medium of culture: Lysogeny broth
growth phase: Exponential
|
| Treatment protocol |
The samples for RNA sequencing were treated with “RNAProtect Bacteria Reagent” (QIAGEN) following the manufacturer's instructions.
|
| Growth protocol |
All strains were carried out in 100 ml glass-flasks containing 25 ml of Lysogeny Broth (LB) Lennox (Pronadisa) and routinely cultured at 37 °C with shaking. Overnight cultures of P. aeruginosa were washed and diluted in LB to an OD600 of 0.01. They were incubated up to exponential phase of growth (OD600 = 0.6) and diluted again to an OD600 of 0.01. Then, the cultures were grown to reach the exponential (OD600 = 0.6) or early stationary phase of growth (OD600 = 2.5).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was obtained as describedm in Schuster et al. (2003) using the RNeasy mini kit (QIAGEN).
RNA samples were sequenced at the "Centro Nacional de Análisis Genómico" (CNAG), Barcelona (Spain). Ribosomal RNA was removed using "RiboZero rRNA Removal kit for Bacteria" (CNAG). To generate the libraries, 2 μg of RNA were treated with "TruSeq RNA sample preparation kit" (Illumina) combined with a specific strand labelling using dUTPs (Sultan, M., et al., 2012). The sequencing in 2 x 75 pair-end format with Illumina technology was performed.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
mexR*. Exponential growth phase in LB liquid cultures.
|
| Data processing |
The quality of sequences were analyzed and trimmed using FastQC software.
The alignment of sequences and gene expression quantification were carried out using the "CLC Genomics Workbench" software (QIAGEN) using default parameters for reverse strand specificity. Only reads from “forward” fastq files were used in this step.
The numeric value of gene expression was normalized to Reads Per Kilobase of gene per million Mapped reads (RPKM). Subsequently, a cut-off value of 1 was added to each RPKM (RPKM + 1) in order to minimize the misleading fold change values caused by RPKMs close to 0 (Charles D. Warden et al., 2013).
Genome_build: NC_002516
Supplementary_files_format_and_content: Processed_RPKM.xlsx: Excel file containing RPKM values for all genes, calculated as described. Several IDs, synonyms and annotations per gene are included.
|
| |
|
| Submission date |
Jul 07, 2020 |
| Last update date |
Oct 25, 2021 |
| Contact name |
Juan Carlos Oliveros |
| Organization name |
CNB, CSIC
|
| Street address |
Darwin 3
|
| City |
Cantoblanco |
| State/province |
Madrid |
| ZIP/Postal code |
28049 |
| Country |
Spain |
| |
|
| Platform ID |
GPL18644 |
| Series (1) |
| GSE153996 |
The impaired quorum sensing response of MexAB-OprM efflux pump overexpressing mutants is not due to non-physiological efflux of 3-oxo-C12-HSL |
|
| Relations |
| BioSample |
SAMN15471196 |
| SRA |
SRX8685384 |
