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Status |
Public on Aug 02, 2016 |
Title |
1.WT-H3-K4me3 |
Sample type |
SRA |
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Source name |
yeast cells
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Organism |
Saccharomyces cerevisiae |
Characteristics |
strain: WT H3 library: Histone mutation library 1st ip: H3 2nd ip / input: K4me3
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Growth protocol |
Yeast cells were grown in YPD media at 30ºC with constant shaking to OD 0.6-0.8.
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Extracted molecule |
genomic DNA |
Extraction protocol |
Cells were fixed with 1% formaldehyde for 15 minutes at room temperature with occasional shaking, quenched with 0.125 M glycine for 5 minutes at room temperature with occasional shaking, collected by centrifugation, (4000 g, 5 minutes), washed with cold ddH2O supplemented with EDTA-free protease inhibitors cocktail (Roche) and the pellet was resuspended in buffer Z (1 M sorbitol, 50 mM Tris 7.4, 10 mM β-mercaptoethanol) with zymolyase (Seikagaku) at 0.3 - 1 units per 1 ml of original cell volume. Cells were gently rotated at 30ºC for 25 minutes until > 95% of cells were spheroplasted. Spheroplasts were pelleted (6500 g, 10 minutes) and resuspended in NP buffer (10 mM Tris pH 7.4, 1 M sorbitol, 50 mM NaCl, 5 mM MgCl2, 1 mM CaCl2, and 0.075% NP-40, freshly supplemented with 1 mM β-mercaptoethanol, 500 μM spermidine, and EDTA-free protease inhibitor cocktail) at final concentration of 200 OD/ml. Chromatin was digested with 12.5 units/ml MNase (Worthington) for 20 minutes at 37ºC, and digestion was stopped by removing the tubes into ice and addition of 1 volume of ice cold MNase stop buffer ( 220 mM NaCl, 0.2% SDS, 0.2% DOX, 10 mM EDTA, 2%,Triton X-100, EDTA-free protease inhibitor cocktail). Tubes were kept on ice for 10 minutes, vortexed 3 x 10 seconds, centrifuged (16,000 g, 10 minutes, 4ºC), and the supernatant containing the nucleosomes was removed to a fresh tube. See attached paper - http://biorxiv.org/content/early/2016/06/27/060962
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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 |
Pair-end reads were mapped to the yeast genome (sacCer3) using bowtie2 with maximal fragment size of 1000bp. We treated duplicate fragments as potential PCR artifacts. We thus treated the set of unique fragments found as the read-set. We defined mononucleosome fragments as these shorter than 220bp. We used the nucleosome location atlas defined by Weiner et al(Weiner et al., 2015). We measured nucleosome coverage by counting the number of fragments overlapping a window of size 50bp around the center of the nucleosome. Genome_build: sacCer3 Supplementary_files_format_and_content: csv file containing read counts for each nucleosome location in each combinatorial ChIP sample
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Submission date |
Jul 11, 2016 |
Last update date |
May 15, 2019 |
Contact name |
Nir Friedman |
E-mail(s) |
nir@cs.huji.ac.il
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Organization name |
The Hebrew University of Jerusalem
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Department |
School of Engineering
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Street address |
Givat Ram
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City |
Jerusalem |
ZIP/Postal code |
9190401 |
Country |
Israel |
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Platform ID |
GPL19756 |
Series (1) |
GSE84240 |
Elucidating Combinatorial Chromatin States at Single-Nucleosome Resolution |
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Relations |
BioSample |
SAMN05371624 |
SRA |
SRX1924855 |
Supplementary data files not provided |
SRA Run Selector |
Raw data are available in SRA |
Processed data are available on Series record |
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