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| Status |
Public on Oct 18, 2013 |
| Title |
GM12892_H3K36me3_1 |
| Sample type |
SRA |
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| Source name |
Lymphoblastoid Cell Lines
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| Organism |
Homo sapiens |
| Characteristics |
cell line: 12892
antibody: H3K36me3
replicate: 1
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| Treatment protocol |
For ChIP-Seq, cross linking was performed in 1% formaldehyde for 10 minutes at room temperature.
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| Growth protocol |
Biological replicates were grown in separate batches and at separate times. Cells were grown to a density of 0.6-0.8 x 10^6/mL in 15% fetal bovine serum. RNA was obtained from lymphoblastoid cell lines (LCLs) of the same growth as used for the ChIP-seq experiments.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
For ChIP-Seq, nuclear lysates were sonicated using a Branson 250 Sonifier (power setting 2, 100% duty cycle for 7 x 30-s intervals). Clarified lysates corresponding to 20 million cells were treated with 1-5ug of antibody (Table S4) coupled to Protein G Dynabeads (Invitrogen #10003D, New York). The protein-DNA complexes were washed with RIPA buffer and eluted in 1% SDS TE at 65°C. An aliquot of clarified lysate was reserved as “input DNA” and handled in parallel with the immunoprecipitated samples following elution from Dynabeads. For RNA-Seq, Total RNA was extracted using Trizol (Lifetechnologies, Grand Island, NY) reagent according to the manufacturer’s instructions, then purified using the Qiagen RNeasy kit (Qiagen, Valencia, CA) and treated with RNAse-free DNase (Qiagen). RNA integrity was checked on a Bioanalyzer (Agilent, Santa Clara CA) and only RNA with an RNA integrity number (RIN) of > 9.5was used for subsequent library construction. Purified total RNA was depleted of ribosomal RNA using the Ribo-Zero Magnetic Gold Kit (Human/Mouse/Rat) (Epicentre Biotechnologies, Madison, WI).
ChIP DNA sequencing libraries were generated according to Illumina DNA Tru-Seq DNA Sample Preparation Kit Instructions (Illumina Part # FC-121-2001, San Diego, CA). For RNA-Seq, stranded libraries were prepared following a dUTP protocol. Briefly, ~100 ng of rRNA depleted RNA were fragmented with 10 x fragmentation buffer (Lifetechnologies, #AM8740) for 2 min at 70 °C. First-strand cDNA synthesis was primed with random hexamers. Actinomycin D was added to reduce anti-sense artifacts. For second-strand synthesis dTTP was replaced with dUTP. The cDNA was endrepaired, A-tailed and Illumina TruSeq adapters were added. After size-selection on an agarose gel the second strand was eliminated by digestion with AmpErase Uracil Nglycosylase (UNG) (Lifetechnologies, N8080096) and after PCR amplification samples were sequenced on the Illumina Hi-Seq 2000.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
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| Data processing |
Personal genomes were created by adding the SNPs of each individual into hg19. SNPs were obtained from the 1000 Genomes Project and from Chen et al 2012
ChIP-seq reads were aligned to personal genomes with BWA 0.6.1 (options -q 20 and the rest set to defaults)
Peaks were called using MACS2. Genome-wide signal tracks (bigWig) were generated using wiggler.
RNA-seq reads were mapped to personal transcriptomes using TOPHAT 2.0.4.
RNA-seq reads overlapping exons (Gencode v10) were obtained using HT-Seq 0.5.4
Genome_build: hg19
Supplementary_files_format_and_content: Peaks are in the narrowPeak format. bigWig files were generated using wiggler. Abundance measurements from the RNA-seq data are presented in a matrix (geneCount_mat.txt), where each row is a gene and each column is a cell line. The values in the matrix are the total counts (from HT-Seq) across both RNA-seq replicates for that cell line.
Samples represent the individual replicates. The combine data are on the Series record.
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| Submission date |
Sep 18, 2013 |
| Last update date |
May 15, 2019 |
| Contact name |
Maya Kasowski |
| E-mail(s) |
kasowski@stanford.edu
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| Phone |
610-283-5618
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| Organization name |
Stanford University
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| Street address |
300 Pasteur Drive
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| City |
Stanford |
| ZIP/Postal code |
94305 |
| Country |
USA |
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| Platform ID |
GPL11154 |
| Series (1) |
| GSE50893 |
Extensive Variation in Chromatin States Across Humans |
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| Relations |
| BioSample |
SAMN02359349 |
| SRA |
SRX356532 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1233964_SNYDER_HG19_GM12892_H3K36ME3_1.norm5.rawsignal.bw |
1.1 Gb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
| Processed data provided as supplementary file |
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