|
| Status |
Public on Jun 18, 2014 |
| Title |
Wild-type (MG1655) T0 RP rep 1 |
| Sample type |
SRA |
| |
|
| Source name |
Aerobic culture
|
| Organism |
Escherichia coli |
| Characteristics |
strain: MG1655
ethanol treatment: no ethanol exposure
ribosome-protected: Yes
|
| Treatment protocol |
Ethanol was added to 40 g/L once A600 of cultures reached 0.3. Samples were taken prior to ethanol addition (T0), 10 min after addition (T1), and 70 min after addition (T2). Cells were harvested rapidly onto 0.2 µm filters by vacuum then flash-frozen in liquid nitrogen to arrest ribosomes. Ribosome footprinting samples were subjected to micrococcal nuclease digestion followed by monosome isolation by sucrose gradient centrifugation.
|
| Growth protocol |
Cultures were grown aerobically at 37 °C in 1 L volumes of M9 minimal medium, supplemented with MgSO4 (1 mM), CaCl2 (0.1 mM), and glucose (10 g/L) in vigorously shaken (225 rpm) Fernbach flasks.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted from samples using hot phenol. The integrity of total RNA was determined from agarose gels.
Library construction was performed as described in Oh et al. 2011, Cell 147(6):1295-1308 (PMCID: PMC3277850). Briefly, fragments of total mRNA or ribosome protected mRNA fragments were size selected via gel purification, and ligated to a 5' adenylated DNA oligo. After reverse transcription, the single stranded DNA was circularized and amplified by PCR.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
This sample represents ribosome footprints from the indicated cells
|
| Data processing |
Basecalls performed using CASAVA version 1.8.2.
Raw reads were trimmed by two nt from the 5' end (to remove any non-templated nts added by reverse transcriptase), and were then mapped using the Burrows-Wheeler Aligner to the Escherichia coli MG1655 genome (NCBI accession NC_000913).
Signals mapping to non-coding RNA regions were removed from the datasets (see supplementary file).
Each dataset was normalized to reads per million per position prior to further analysis.
For metagene analyses (gene-segment analyses and codon-type analyses), pseudogenes and genes not represented in one or more datasets were excluded, leaving 3048 genes in the "all genes" dataset (see supplementary file for list of genes in "all genes", "low ribosome occupancy", and "high ribosome occupancy" gene sets)
Genome_build: NC_000913.2
Supplementary_files_format_and_content: Processed data files are WIG files with non-coding RNAs removed, normalized to reads per million per position. Those with a "-" as the final character in the file name cover the minus strand of the genome; those with a "+" as the final character in the file name cover the plus strand of the genome.
Supplementary_files_format_and_content: Gene_lists_GEO.xls; The supplementary file contains gene lists used in data analysis.
|
| |
|
| Submission date |
Mar 31, 2014 |
| Last update date |
May 15, 2019 |
| Contact name |
Rembrandt Haft |
| Organization name |
University of Wisconsin
|
| Department |
Great Lakes Bioenergy Research Center
|
| Lab |
Landick Lab
|
| Street address |
1550 Linden Dr. Room 5445
|
| City |
Madison |
| State/province |
WI |
| ZIP/Postal code |
53703 |
| Country |
USA |
| |
|
| Platform ID |
GPL14548 |
| Series (2) |
| GSE56372 |
Correcting Direct Effects of Ethanol on Translation and Transcription Machinery Confers Ethanol Tolerance in Bacteria [Ribosome_Profiling] |
| GSE56408 |
Correcting Direct Effects of Ethanol on Translation and Transcription Machinery Confers Ethanol Tolerance in Bacteria |
|
| Relations |
| BioSample |
SAMN02713613 |
| SRA |
SRX505567 |
