|
| Status |
Public on Mar 28, 2018 |
| Title |
Bacillus_subtilis_pnpA_Rend_seq_LB_25s_frag |
| Sample type |
SRA |
| |
|
| Source name |
Bacteria
|
| Organism |
Bacillus subtilis subsp. subtilis str. 168 |
| Characteristics |
strain: subspecies 168, pnpA::lox72
growth medium: LB
|
| Growth protocol |
Overnight cultures were diluted to OD590 0.0003 in 20 mL fresh media. The culture was kept in a 125 mL flask at 30C with aeration (200 rpm) until OD590 reached 0.3.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
5 mL of cell culture was added to 5 mL of cold (-30C) methanol, mixed by inversion and spun down at 3000 rcf for 10 min at 4C. The supernatant was decanted and the cell pellet frozen at -80C. RNA was extracted using the RNAeasy kit (QIAGEN).
Ribosomal RNA was depleted using the MICROBExpress kit (Thermo Fisher). The resulting RNA was fragmented for 25 s at 95C using RNA fragmentation reagents (Thermo Fisher, AM 8740). Fragments in the 15 to 45 nt range were selected, dephosphorylated at the 3’ end and ligated to a 5’ adenylated DNA oligo. After reverse transcription, the single stranded DNA was circularized, and PCR amplified.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
mRNA
|
| Data processing |
Sequence reads were trimmed for adaptor sequences.
Trimmed reads were aligned to NC_000964.3 using Bowtie v1.0.1 with options -v 1 -k 1. To deal with non-template addition during reverse transcription, reads with a mismatch at their 5' end had their 5' end re-assigned to the immediate next downstream position.The 5' and 3' ends of mapped reads between 15 and 45 nt in sizes were added separately at genomic positions to generate the wig file (thus leading to 4 wig files: 3’ forward, 3’ reverse, 5’ forward, 5’ reverse).
NC_000964.3
Base calls performed using Casava version 1.7.
Supplementary_files_format_and_content: wiggle file with two columns: first column containing chromosome positions and second column containing the number of reads mapped to the position (see publication for details).
|
| |
|
| Submission date |
Jan 30, 2018 |
| Last update date |
Mar 28, 2018 |
| Contact name |
Jean-Benoit Lalanne |
| E-mail(s) |
lalannej@uw.edu
|
| Organization name |
University of Washington
|
| Department |
Genome Sciences
|
| Lab |
Jay Shendure
|
| Street address |
3720 15th Ave NE
|
| City |
Seattle |
| State/province |
WA |
| ZIP/Postal code |
98195 |
| Country |
USA |
| |
|
| Platform ID |
GPL15205 |
| Series (1) |
| GSE95211 |
Evolutionary Convergence of Pathway-specific Enzyme Expression Stoichiometry |
|
| Relations |
| BioSample |
SAMN08437337 |
| SRA |
SRX3630292 |
