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Status |
Public on Mar 30, 2017 |
Title |
ChIP-seq_Spt5-deg_DMSO_Pol2_r2 |
Sample type |
SRA |
|
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Source name |
Strain_FWP486_genotype_Spt5-deg
|
Organism |
Schizosaccharomyces pombe |
Characteristics |
growth_medium: EMM treatment: DMSO hours_of_treatment: 4.5 spike_in: 10% S. cerevisiae chromatin demultiplex_code: AGGTCAGTT spt5_gene_status: nmt81-Spt5-V5-IAA17 antibody: 8WG16 antibody_target: Pol2 replicate: 2
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Treatment protocol |
Spt5 depletion experiments were performed by adding NAA (0.5mM in DMSO, napthaleneacetic acid, Sigma, N0640) and thiamine (100nM, thiamine hydrochloride, Sigma, T4625), to cells grown to a density of 10^7 cells/ml (O.D.600 ~ 0.5). Cultures were incubated with shaking at 30°C for 4.5 hours.
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Growth protocol |
S. pombe strains were grown in EMM complete media at 30°C.
|
Extracted molecule |
genomic DNA |
Extraction protocol |
Samples were prepared as described in (PMID:24100010) ChIP-seq libraires were prepared as described in (PMID:24100010). For the spike-in control, S. cerevisiae chromatin was added to S. pombe chromatin before immunoprecipitation with the relevant antibodies. S. cerevisiae chromatin corresponding to 10% of S. pombe chromatin, as estimated by Bradford assay (Bio-Rad), was used. Libraries were sequenced on the Illumina NextSeq 500 platform at the Harvard Bauer Core facility.
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Library strategy |
ChIP-Seq |
Library source |
genomic |
Library selection |
ChIP |
Instrument model |
Illumina NextSeq 500 |
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Data processing |
Demultiplexing, 5' barcode removal, and 3' trimming of initial 75bp sequence to 60bp resulted in the attached FASTQs. Alignment using Bowtie 1.1.1 (parameter -m 1 for unique mapping) to a concatenated S. pombe and S. cerevisiae genome. Split resulting BAMs by organism, then counted reads to produce ratios for spike-in normalization. Determined cross-correlations via SPP which recommended tag shift of ~180bp for all ChIP samples. bedGraph track files produced for S. pombe of 180bp tag-shifted density with bandwidth 20, sampled every 10bp, spike-in-normalized. Genome_build: S. pombe ASM294v2 concatenated to S. cerevisiae R64-1-1 Supplementary_files_format_and_content: bedGraph track files for S. pombe tag-shifted density with bandwidth 20, sampled every 10bp, spike-in-normalized.
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Submission date |
Jan 18, 2017 |
Last update date |
May 15, 2019 |
Contact name |
Peter J Park |
E-mail(s) |
peter_park@harvard.edu
|
Phone |
617-432-7373
|
Organization name |
Harvard Medical School
|
Department |
Center for Biomedical Informatics
|
Street address |
10 Shattuck St
|
City |
Boston |
State/province |
MA |
ZIP/Postal code |
02115 |
Country |
USA |
|
|
Platform ID |
GPL20584 |
Series (2) |
GSE85182 |
Genome-wide analyses reveal widespread roles for Spt5 in sense and antisense transcription |
GSE93803 |
Genome-wide analyses reveal widespread roles for Spt5 in sense and antisense transcription [ChIP-seq_round2] |
|
Relations |
BioSample |
SAMN06239107 |
SRA |
SRX2501576 |