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| Status |
Public on Apr 14, 2021 |
| Title |
J_R1 |
| Sample type |
SRA |
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| Source name |
U2OS cells
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| Organism |
Homo sapiens |
| Characteristics |
cell line: U2OS
overexpressed protein: DNAJC9-MYC-Flag J mutant
spike-in: YES
antibodies: FLAG: Sigma, F7425; H2Av: Active Motif, 39715
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| Treatment protocol |
Protein expression was induced in media with Tetracycline (2 µg/ml, 48 hrs) in DMEM, 10 % FBS and 1 % P/S.
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| Growth protocol |
U2OS were grown in DMEM with 10 % heat-inactivated FBS and 1 % penicillin/streptomycin under Blasticidin (5 µg/ml) selection and additionally Hygromycin B (100 µg/ml) selection for cell lines with pcDNA5-FRT-TO integration at 37 °C with 5 % CO2. Drosophila S2 cells (male) were grown in M3+BPYE media: Shields and Sang M3 Insect Medium, KHCO3, yeast extract, bactopeptone, 10 % heat-inactivated FBS and 1X penicillin/streptomycin at 25 °C with 5 % CO2.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Cells were processed using the truChIP Chromatin Shearing Kit (Covaris, 520127) following manufacturer’s instructions. 20 million nuclei were sonicated in 1mL tubes using a Covaris M220 to obtain chromatin frgament size of 250-500bp. Sonicated chromatin was centrifuged at 14,000 rpm at 4ºC for 10 minutes and the supernatant was isolated for subsequent steps. In parallel, Drosophila S2 cells were fixed, lysed and sonicated as described above. U2OS input chromatin was mixed with Drosophila S2 chromatin (1.5% of total chromatin) after sonication. Chromatin associated to DNAJC9-MYC-Flag was obtained by incubating sonicated chromatin with anti FLAG antibody.
Libraries were costructed using the KAPA Hyperprep kit following manufacturer’s instruction, except that 1.25 µM of Illumina compatible indexed adapters (Pentabase) were ligated to A-tailed DNA.
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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 |
Illumina RTAv2 software was used for basecalling
Raw reads were trimmed using Trim Galore! (Galaxy version 0.4.2) to remove low-quality reads and adaptor sequence.
Libraries were mapped to the hg38 and dm3 genome assemblies using Bowtie2 (Galaxy version 2.2.6.2).
Duplicates and reads mapping to the Broad Institute's blacklist (Encode Project Consortium, 2012) were discarded. Remaining reads were extended by 250 bp for downstream analyses. Replicates were merged for plotting and downstream analyses.
Genome_build: hg38 and dm3
Supplementary_files_format_and_content: Bedgraph files contain either RPM, RRPM or RRPM, n+1, log-transformed read counts for merged replicates (R1+R2) of ChIP-seq datasets from 10 Kb windows with a 5 Kb step.
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| Submission date |
Jul 14, 2020 |
| Last update date |
Apr 15, 2021 |
| Contact name |
Anja Groth |
| E-mail(s) |
anja.groth@cpr.ku.dk
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| Organization name |
Novo Nordisk Foundation Center for Protein Research
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| Street address |
Blegdamsvej 3B
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| City |
Copenhagen |
| ZIP/Postal code |
2200 |
| Country |
Denmark |
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| Platform ID |
GPL18573 |
| Series (1) |
| GSE154445 |
DNAJC9 Integrates Heat Shock Molecular Chaperones into the Histone Chaperone Network |
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| Relations |
| BioSample |
SAMN15537645 |
| SRA |
SRX8733219 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4671245_J_RPM_merged.bedGraph.gz |
4.7 Mb |
(ftp)(http) |
BEDGRAPH |
| GSM4671245_J_RRPM_log_n+1_merged.bedGraph.gz |
5.2 Mb |
(ftp)(http) |
BEDGRAPH |
| GSM4671245_J_RRPM_merged.bedGraph.gz |
4.8 Mb |
(ftp)(http) |
BEDGRAPH |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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