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| Status |
Public on Jan 01, 2017 |
| Title |
HfqEx2 |
| Sample type |
SRA |
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| Source name |
Bacteria
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| Organism |
Pseudomonas putida |
| Characteristics |
strain/genotype: KT2440 Δhfq
growth phase: exponential
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| Treatment protocol |
For all experiments the overnight culture of P. putida was diluted to starting OD600 0.1 in 100 ml of LB. Cells were harvested at three time points on the growth curve: mid-exponential (OD600~0.5), transition (OD600~2.5), and stationary phase (twice doubling time after the transition phase). For size-selected library KB1 three biological replicates of wt and Δhfq strains were grown.
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| Growth protocol |
P. putida KT2440 was grown at 30 °C, 250 rpm and routinely cultured in LB medium according to standard protocols
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| Extracted molecule |
total RNA |
| Extraction protocol |
RNA extraction for cDNA library KB1 preparations was performed as previously described (Gómez-Lozano et al., 2012). Briefly, 10 ml of harvested culture was mixed with 0.2 volumes of STOP solution (95% [v/v] ethanol, 5% [v/v] phenol). Cells were centrifuged, 1 mL of Trizol (Invitrogen) was added and the samples were snap frozen. Total RNA was extracted and treated with DNase I (Fermentas). Total RNA integrity and quality were validated by Bioanalyzer (Agilent).
For KB1 cDNA library, RNA was size-selected (up to 500 nt) as described previously (Gómez-Lozano et al., 2012) using 10% polyacrylamide-urea gels containing urea (Bio-Rad) with some changes. Sample was depleted of rRNA using MICROBExpress Kit (Ambion) and treated with Tobacco Acid Pyrophosphatase TAP (Epicentre). Following the fragmentation with RNaseIII, libraries were prepared using TruSeq Small RNA Sample Preparation Kit (Illumina).
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
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| Description |
5 A
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| Data processing |
The RNA-seq data was trimmed by Beckman
RNA-seq data was analysed with open source software Rockhopper (version 2.0.3) (McClure et al., 2013)
Differential gene and sRNA expression analysis were carried out with the webserver T-REx (de Jong et al., 2015) using the RPKM values generated in the Rockhopper analysis. Differential expression of genes was considered significant with a fold change ≥ 2 and adjusted p-value ≤ 0.05
Genome_build: GenBank accession no. NC_002947.3
Supplementary_files_format_and_content: One file with all the processed results in excel: Raw counts, Normalized counts, RPKM, Expression values, q-values
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| Submission date |
Aug 13, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Klara Bojanovic |
| E-mail(s) |
klara.bojanovic@gmail.com
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| Phone |
004524653766
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| Organization name |
Technical University of Denmark
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| Department |
Novo Nordisk Foundation Center for Biosustainability
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| Street address |
Building 220, Kemitorvet
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| City |
Kgs. Lyngby |
| ZIP/Postal code |
2800 |
| Country |
Denmark |
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| Platform ID |
GPL19659 |
| Series (1) |
| GSE85578 |
RNA-sequencing data from Pseudomonas putida KT2440 wild-type and Δhfq mutant |
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| Relations |
| BioSample |
SAMN05572235 |
| SRA |
SRX2020717 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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