|
| Status |
Public on Feb 15, 2017 |
| Title |
Eco_TolC_30min_control_A |
| Sample type |
SRA |
| |
|
| Source name |
planktonic cells
|
| Organism |
Escherichia coli |
| Characteristics |
treatment: none
genotype: TolC defective (TolC-)
time point (minutes): 30
|
| Treatment protocol |
An o/n culture of E. coli TolC was diluted to an OD600 of 0.1 and supplied with a final concentration of 0.5 µg/ml Carolacton in MeOH. Controls received the same volume MeOH. Three early time-points (t) were chosen to be analyzed by RNA-seq. Prior to addition of Carolacton (t0), t5, t15 and t30 min after addition of Carolacton. At each point, 5 ml of culture were mixed with an equal volume RNAprotect (Qiagen), incubated at room temperature (RT) for 5 min, the cells harvested by centrifugation, and the pellet frozen and stored at -80°C.
|
| Growth protocol |
E. coli TolC cells were grown o/n in LB medium at 37°C and 200 rpm.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total cellular RNAs were extracted using the RNeasy Mini Kit (Qiagen) according to the manufacturers protocol. Residual DNA in the fractions was digested by addition of 27 Kunitz units Dnase I (Qiagen), and absence of DNA verified by Polymerase chain reaction (PCR). mRNA enrichment was conducted using the Ribo-Zero kit for Gram-negative bacteria (epicentre). RNA quality control of the RNA extracts, as well as the successful mRNA enrichment, was conducted by using an Agilent 2100 Bioanalyzer.
Libraries of RNA fractions were constructed with the ScriptSeq V2 RNA-Seq Library Prep Kit (Illumina) according to the manufacturers guidelines.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Data processing |
Sequenced reads were trimmed for adaptor sequences using FastQC v0.11.2.
Raw read counts of transcripts were obtained by aligning samples 1-14 to the reference genome and counting mapped reads per feature using Rockhopper v2.0.3.
Genome_build: CP018801 (https://www.ncbi.nlm.nih.gov/nuccore/CP018801)
Supplementary_files_format_and_content: The tab-delimited text files include raw read values for each time-point for both biol. replicates of control and Carolacton-treated samples as obtained from Rockhopper. A FASTA file on the series record contains the transcript sequences.
|
| |
|
| Submission date |
Jan 04, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Jannik Donner |
| E-mail(s) |
[email protected]
|
| Organization name |
Helmholtz-Center for Infection Research
|
| Street address |
Inhoffenstr. 7
|
| City |
Braunschweig |
| ZIP/Postal code |
38124 |
| Country |
Germany |
| |
|
| Platform ID |
GPL18133 |
| Series (1) |
| GSE93125 |
Transcriptomal changes of Escherichia coli TolC grown with the biofilm inhibitor Carolacton |
|
| Relations |
| BioSample |
SAMN06200382 |
| SRA |
SRX2462349 |
