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Status |
Public on Jan 25, 2024 |
Title |
Human_ZSCAN4_Rabbit_Ab_DUX4_plus |
Sample type |
SRA |
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Source name |
Embryonic cells
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Organism |
Homo sapiens |
Characteristics |
tissue: Embryonic cells cell line: ES cells (SEES3) cell type: Pluripotent stem cells genotype: Male antibody: Human_Zscan4_Rabbit_Ab treatment: DUX4_plus replication: Technical replicate 2
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Extracted molecule |
genomic DNA |
Extraction protocol |
Cells were cross-linked with 2 mM disuccinimidyl glutarate in PBS/1 mM MgCl2 for 40 min at room temperature followed by 1% formaldehyde in PBS for 15 min at room temperature. The reaction was stopped by 125 mM glycine. The cells were washed with PBS and stored at –80°C prior to use. The cells were lysed in Lysis buffer (10 mM Tris-HCl, pH8.0, 1 mM EDTA, 0.5 mM EGTA, 100 mM NaCl, 0.1% Na-Deoxycholate, 0.5% N-lauroyl sarcosine) containing proteinase inhibitor cocktail (Roche). Sonication was conducted with the Handy sonicator UR-21P (Tomy) to generate DNA fragments of approximately 150-450 bp. The sonicated lysates from approximately 2 x 106 cells were diluted in ChIP dilution buffer (10 mM HEPES, pH 7.4, 50 mM NaCl, 1% IGEPAL-CA-630, 10% glycerol) containing proteinase inhibitor cocktail and incubated overnight at 4°C with 50 µl of protein G magnetic beads (Invitrogen) that were preincubated with ~2 µg of mouse Zscan4 antibodies. The precipitants were washed once with high salt wash buffer (20 mM Tris-HCl, pH8.0, 400 mM NaCl, 2 mM EDTA, 0.1% SDS, 0.2% Triton-X), three times with LiCl wash buffer (10 mM Tris-HCl, pH 8.0, 1 mM EDTA, 250 mM LiCl, 1% NP40, 1% Na-deoxycholate) and once with 10 mM Tris-HCl, pH 8.0, 5 mM EDTA, 10 mM NaCl. Bound chromatin was eluted in elution buffer (90 mM NaHCO3, 1% SDS), followed by RNase treatment at 37°C for 30 min and crosslink reversal with decross-linking mixture (2 M NaCl, 0.1 M EDTA, 0.4 M Tris-HCl, pH 6.8) containing proteinase K at 65°C for 3 h. DNA was purified by a phenol-chloroform-isoamylalcohol extraction and ethanol precipitation. ChIP DNA libraries were prepared with the NEBNext ChIP-Seq Library Prep Kit for Illumina (NEB)
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Library strategy |
ChIP-Seq |
Library source |
genomic |
Library selection |
ChIP |
Instrument model |
Illumina HiSeq 2500 |
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Data processing |
Fastq files were processed and analyzed on the Biowardrobe platform Sequence reads were aligned using bowtie (version 1.2.0) MACS2 (version 2.1.1.20160309) was used to estimate fragment size and to find islands of enrichment with q-value threshold less than 0.2 To visualize the data, the data were uploaded to UCSC genome browser MAnorm was used for comparison of ChIP peaks Heat maps and average signal profiles were generated by Easeq Assembly: mm10, hg19 Supplementary files format and content: bigwig
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Submission date |
Sep 20, 2023 |
Last update date |
Jan 25, 2024 |
Contact name |
Tomohiko Akiyama |
E-mail(s) |
akiakiakit@hotmail.com
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Phone |
0457872596
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Organization name |
Yokohama city university
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Department |
School of medicine
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Street address |
3-9 Fuku-ura
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City |
Yokohama |
State/province |
Kanagawa |
ZIP/Postal code |
236-0004 |
Country |
Japan |
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Platform ID |
GPL16791 |
Series (2) |
GSE243626 |
ZSCAN4-binding motif - TGCACAC is conserved and enriched in CA/TG microsatellites in both mouse and human genomes [ChIP-Seq] |
GSE243628 |
ZSCAN4-binding motif - TGCACAC is conserved and enriched in CA/TG microsatellites in both mouse and human genomes |
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Relations |
BioSample |
SAMN37480942 |
SRA |
SRX21841556 |
Supplementary file |
Size |
Download |
File type/resource |
GSM7792026_Human_ZSCAN4_Rabbit_Ab_DUX4_plus.bw |
116.1 Mb |
(ftp)(http) |
BW |
SRA Run Selector |
Raw data are available in SRA |
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