|
| Status |
Public on May 01, 2018 |
| Title |
Hfq_stationary_Rif A |
| Sample type |
SRA |
| |
|
| Source name |
Bacterial cells grown to OD 2.0
|
| Organism |
Pseudomonas aeruginosa PAO1 |
| Characteristics |
optical density: OD 2.0
genotype: Hfq-VSVG FRT
treatment protocol: sample taken 30 minutes after treatment with 150 ug/mL rifampicin
chip antibody: Agarose-immobilized anti-VSVG antibody (Sigma-Aldrich, catalog number A1970, monoclonal antibody clone P5D4)
|
| Growth protocol |
Bacterial cultures were started from single colonies, grown overnight, and back-diluted in LB media
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
DNA associated with proteins of interest were immunoprecipitated from lysates with anti-VSVG monocolonal antibody
Libraries were prepared using NEBNext Ultra II DNA Library Prep Kit for Illumina (NEB) following manufacturer’s instructions, bead-based size selection of DNA fragments was used in a subset of samples, see SAMPLES section
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Description |
Immunoprecipitated DNA was size-selected using AMPure beads during library construction
Hfq_stat_Rif_AllPeaks.bed
R1 raw file checksum: 9a44a73c0adb1ef52a6c220dee0b0ddb
R2 raw file checksum: 37e04d13b8c40ccde6b3f9ff9ec5f32e
|
| Data processing |
Paired ChIP-Seq reads corresponding to fragment sizes of either 200 or 400 bp (see SAMPLES section) were aligned to the PAO1 genome using bowtie2 (version 2.3.1) allowing 1 mismatch in a 28 bp seed region with no discordant or unpaired alignments.
The program samtools (version 1.3.1) was used to extract read 1 from each mapped read pair.
For Hfq experiments (samples 1-4): Peaks were identified using QuEST (version 2.4; Valouev et al., 2008) using control data from stationary-phase wild-type cells (sample 5) with the following settings: KDE = 50, seeding fold enrichment = 1.25, extension fold enrichment = 1.5, ChIP to background fold enrichment = 2
For Crc experiments (samples 6 and 7): Peaks were identified using QuEST (version 2.4; Valouev et al., 2008) using control data from log-phase wild-type cells (sample 8) with the following settings: KDE = 30, seeding fold enrichment = 1.25, extension fold enrichment = 1.5, ChIP to background fold enrichment = 2; only peaks with greater than 3x enrichment were considered.
Bed files for each experimental replicate were created by QuEST and the final processed bed files were generated by identifying the largest overlapping peak region which statisfy the following conditions: (1) peak regions must have a positive peak shift, (2) peak regions must have a positive strand correlation, (3) peak regions must have a q-value of less than 0.01, and (4) peak regions must be shared by two or more experimental replicates.
Genome_build: NC_002516.2
Supplementary_files_format_and_content: wig files were generated using QuEST; Scores represent normalized smoothed read density
Supplementary_files_format_and_content: bed intervals reflect regions with significant (as defined above) enrichment of reads in the experimental condition in comparision to the mock control condition.
|
| |
|
| Submission date |
Feb 23, 2018 |
| Last update date |
May 01, 2018 |
| Contact name |
Simon L Dove |
| E-mail(s) |
[email protected]
|
| Organization name |
Boston Children's Hospital and Harvard Medical School
|
| Department |
Division of Infectious Diseases
|
| Lab |
Dove Laboratory
|
| Street address |
300 Longwood Ave
|
| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02115 |
| Country |
USA |
| |
|
| Platform ID |
GPL18782 |
| Series (1) |
| GSE111037 |
Co-transcriptional association of Hfq and Crc with the Pseudomonas aeruginosa PAO1 genome |
|
| Relations |
| BioSample |
SAMN08284316 |
| SRA |
SRX3528049 |
| BioSample |
SAMN08583740 |
