|
| Status |
Public on Aug 01, 2022 |
| Title |
wild-type |
| Sample type |
SRA |
| |
|
| Source name |
cell cultures
|
| Organism |
Escherichia coli str. K-12 substr. MG1655 |
| Characteristics |
genotype/variation: wild-type
plasmids: IRAGP_reporter
|
| Growth protocol |
Cultures with the IRAGP_reporter plasmid were harvested at OD600 = 0.3 at 37 C by filtration.
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
Cell pellets were cryogenically pulverized in a Spex 6870 freezer mill; 5 cycles of 1 min at 5 Hz with 1 min cooling. The lysis buffer contained Cm to arrest transation. Lysates were clarified and ribosomes were pelleted prior to digestion of mRNA with MNase. After digestion, 70S monosomes were purified over a sucrose gradient.
Ribosome footprints and total RNA fragments 15-45 nt in length were PAGE purified, ligated to a linker using RNA ligase T2, converted to DNA using RT, circularized, and PCR amplified.
|
| |
|
| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Data processing |
Library strategy: Ribo-seq
The linker was trimmed using Skewer. Reads were aligned using Bowtie and allowing two mistmatches. Reads mapping to rRNA or tRNA sequences were discarded. The remaining reads were mapped to the E. coli K-12 MG1655 genome version 2. Separately, the reads were mapped to the IRAGP_reporter plasmid sequence. In the WIG files, rpm values include the fraction of reporter reads in all the uniquely mapped reads, both the chromosome and the plasmid.
Genome_build: NC_000913.2 and separately the IRAGP_reporter plasmid
Supplementary_files_format_and_content: WIG files of ribosome occupancy, assigned to the 3'-end of reads, normalized by the total number of reads in the library.
|
| |
|
| Submission date |
Jul 07, 2021 |
| Last update date |
Aug 01, 2022 |
| Contact name |
Allen R Buskirk |
| E-mail(s) |
[email protected]
|
| Organization name |
Johns Hopkins University School of Medicine
|
| Department |
Molecular Biology and Genetics
|
| Street address |
725 N. Wolfe St
|
| City |
Baltimore |
| State/province |
MD |
| ZIP/Postal code |
21205 |
| Country |
USA |
| |
|
| Platform ID |
GPL18956 |
| Series (1) |
| GSE179691 |
Ribosome collisions in bacteria promote ribosome rescue by triggering mRNA cleavage by SmrB |
|
| Relations |
| BioSample |
SAMN20110714 |
| SRA |
SRX11375874 |
