|
| Status |
Public on Apr 04, 2020 |
| Title |
ura3FLAGlog_IP |
| Sample type |
SRA |
| |
|
| Source name |
archaeal cells
|
| Organism |
Halobacterium salinarum |
| Characteristics |
strain: AKS113
genotype: [delta]ura3 cdrL::FLAG
growth phase: logarithmic
epitope tag: FLAG
antibody: anti-FLAG (Abcam ab1162) anti-rabbit monoclonal antibody
|
| Treatment protocol |
Cells were crosslinked and immunopreciptated with FLAG antibody as described in Wilbanks et al., 2012 (doi: 10.1093/nar/gks063) with the following exceptions: cultures were crosslinked and immuniprecipitated as described (Wilbanks et al., 2012) with the following exceptions: cultures were crosslinked with 1% formaldehyde for 30 minutes at room temperature; immuoprecipitations were conducted using Dynabead magnetic beads (Thermo-Fisher product 10002D) conjugated with anti-FLAG (Abcam ab1162) anti-rabbit monoclonal antibody at 1:250 dilution. DNA concentration was determined by Nanodrop (Thermo Scientific).
|
| Growth protocol |
Triplicate cultures of strains AKS113 (CdrL tagged at the C-terminus with the FLAG epitope, Supplementary Table 4) and ∆ura3 control strain were grown until stationary phase and subcultured in rich media supplemented with 50 µg/ml uracil. At mid-log phase (OD600 ~0.15) and early stationary phase (OD600 ~1.8), cultures were crosslinked and immunoprecipitated.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Libraries were constructed using the KAPA Hyper Prep kit and Illumina TruSeq adapters according to manufactuer's instructions. DNA library quality was assessed by Bioanalyzer using a High Sensitivity DNA chip (Agilent). Samples were pooled and run in a single lane on an Illumina HiSeq 4000 (Duke Sequencing and Genomics Technologies core).
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 4000 |
| |
|
| Description |
DNA immunopreciptated control with no epitope tag
HBT-ura3-cdrL-log-pks.bed
|
| Data processing |
50 bp single reads were assessed for quality using FastQC (www.bioinformatics.babraham.ac.uk)
adapter sequences trimmed using Trim Galore (www.bioinformatics.babraham.ac.uk) and Cutadapt
Resultant sequences were aligned to H. salinarum NRC-1 genome (RefSeq: NC_002607.1, NC_002608.1, NC_001869.1) using Bowtie2
Peaks were called using MOSAiCS with arguments: fragment length 200, bin size 200, read capping 0, analysis type IO, background estimate rMOM, signal model 2S, FDR 0.01.
Genome_build: GCF_000006805.1_ASM680v1
Supplementary_files_format_and_content: wig files were generated using the "generateWig()" command in the R MOSAiCs package; bed files were generated from peak calling with MOSAiCs
|
| |
|
| Submission date |
Apr 03, 2020 |
| Last update date |
Apr 05, 2020 |
| Contact name |
Amy K Schmid |
| E-mail(s) |
[email protected]
|
| Organization name |
Duke University
|
| Department |
Biology
|
| Lab |
Schmid
|
| Street address |
125 Science Dr.
|
| City |
Durham |
| State/province |
NC |
| ZIP/Postal code |
27707 |
| Country |
USA |
| |
|
| Platform ID |
GPL28343 |
| Series (1) |
| GSE148065 |
CdrL ChIP-seq data for the paper "CdrS is required for cell division site placement but not elongation in hypersaline-adapted archaea" |
|
| Relations |
| BioSample |
SAMN14534536 |
| SRA |
SRX8053625 |
