|
| Status |
Public on Mar 28, 2017 |
| Title |
B CCB604 (W168 [delta]yacP + pDG-yacP) + IPTG |
| Sample type |
SRA |
| |
|
| Source name |
cell culture
|
| Organism |
Bacillus subtilis |
| Characteristics |
strain: CCB604 (W168 [delta]yacP + pDG-yacP) + IPTG
|
| Growth protocol |
Cultures were grown at 37°C in 2xYT in the presence of appropriate antibiotics (mls or kan) with shaking +/- 1 mM IPTG. Cells were harvested by centrifugation at OD600 = 0.6
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated from 20 mL culture (pelleted and frozen) by the glass beads/phenol method described previously (Bechhofer et al, 2008). RNA samples were treated with RQ DNase Promega (37°C for 20 minutes) to remove potential contaminating chromosomal DNA.
Ribosomal RNA was removed from 5 mg total RNA using RiboZero kit from Epicenter, according to the manufacturer's instructions. Ribosomal RNA depletion and overall RNA quality was analysed by Bioanalyser (Agilent). cDNA libraries were prepared using the Smarter Stranded RNA-Seq Kit (Clontech) with adapters for multiplexing, according to manufacturer's instructions. RNA concentration and quality was checked by Bioanalyser (Agilent).
The 12 samples were normalized to 2nM, multiplexed and denatured at a concentration of 1nM using 0.1N NaOH (5 minutes at room temperature) before dilution to 10pM and loading on a HiSeq2000 flowcell.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
SLX085-02-v4.604p_complete.xls
SLX085-02-v4.604mvs604p.complete.xls
|
| Data processing |
Reads were demultiplexed and adapters trimmed (https://demult.web.pasteur.fr/ and Casava 1.8 version (Consensus Assessment of Sequence and Variance software, Illumina).
Quality control checks were performed before and after trimming by FastQC (http://www.bioinformatics.babraham.ac.uk/projects/fastqc/).
Fifty-nt reads were mapped by Bowtie (Langmead et al, 2009) and yielded about 107 aligned reads per sample.
Data analysis was performed using the R software, Bioconductor (Gentleman et al, 2004) packages including DESeq2 (Anders & Huber, 2010; Love et al, 2014) and the PF2tools package (version 1.2.3) developed at PF2 (Institut Pasteur).
Normalization and differential analysis were carried out according to the DESeq2 model and package (version 1.6.2).
Supplementary_files_format_and_content: Excel files
|
| |
|
| Submission date |
Jan 20, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Ciaran Condon |
| E-mail(s) |
condon@ibpc.fr
|
| Organization name |
CNRS
|
| Department |
UMR8261
|
| Lab |
Institut de Biologie Physico-Chimique
|
| Street address |
13 rue Pierre et Marie Curie
|
| City |
Paris |
| ZIP/Postal code |
75005 |
| Country |
France |
| |
|
| Platform ID |
GPL18561 |
| Series (1) |
| GSE93894 |
RNAseq of ∆yacP (∆rae1) mutant and plasmid complemented strain |
|
| Relations |
| BioSample |
SAMN06246867 |
| SRA |
SRX2508587 |
