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Sample GSM2284155

Query DataSets for GSM2284155

Status

Public on Sep 29, 2016

Title

Input_2

Sample type

SRA

Source name

in vitro transformed human skin fibroblasts

Organism

Homo sapiens

Characteristics

cell type: in vitro transformed human skin fibroblasts
chip antibody: none

Growth protocol

Cells were grown in minimum essential medium (MEM) containing 15% fetal bovine serum (FBS), 2 mM L-glutamine, 100 U/ml penicillin and 100 μg/ml streptomycin, at 37 °C in 5% CO2

Extracted molecule

genomic DNA

Extraction protocol

Cells were harvested with trypsin and ChIP was performed as previously described (Sailaja et al., Methods in molecular biology, 2012).
Library construction was carried out as published (Blecher-Gonen et al., 2013, Nat Prot). Paired end libraries were constructed using standard Illumina protocols, with some modifications. Agencourt AMPure XP (Beckman Coulter) at 0.8x ratio were used to size select out adapter dimers. The Illumina Phusion enzyme was replaced by Kapa HiFi HotStart ready mix (Kapa Biosystems). An Invitrogen SizeSelect R-gel system (Life Technologies) was used to size select following PCR amplification.

Library strategy

ChIP-Seq

Library source

genomic

Library selection

ChIP

Instrument model

Illumina HiSeq 2500

Data processing

Raw reads that passed the quality controls were aligned to human genome assembly hg19 using Bowtie 0.12.7 (Langmead et al. Genome Biol., 2009), taking only uniquely aligned reads with no more than 2 mismatches.
Significant peaks of H3K4me3 and H3K27me3 were extracted using MACS 1.4 (Zhang et al., 2008), using a minimal p-value cutoff of 10E-5 and a fold change for model building of 30.
Peak detection for H1.0 and H1.4 was performed using in house scriots. In short, reads were counted within 50 bp windows. The median was calculated for each three bins and normalized by the sum of the reads. To effectively capture local biases in the genome, the Log2 ratio between the ChIP samples and the input genomic DNA was calculated. Peaks were defined as regions with log2 ratio of ChIP DNA over Input DNA > 2 within six neighbouring windows, and average number of reads > 15.
Genome_build: hg19
Supplementary_files_format_and_content: bed format

Submission date

Aug 18, 2016

Last update date

May 15, 2019

Contact name

Eran Meshorer

E-mail(s)

meshorer@huji.ac.il

Organization name

The Hebrew University of Jerusalem

Department

Genetics