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Sample GSM1652703

Query DataSets for GSM1652703

Status

Public on Nov 09, 2015

Title

Input_SET4_MxEmb_Rep1

Sample type

SRA

Source name

whole worms

Organism

Caenorhabditis elegans

Characteristics

strain: MT14911
developmental stage: mixed embryo
antibody: none (input)

Treatment protocol

Embryos were treated with 2% formaldehyde for 30 minutes, washed with M9, and collected by centifugation and freezing at -80C until extract preparation.

Growth protocol

Embryos were isolated by bleaching gravid adults grown on plates at 20C temperature.

Extracted molecule

genomic DNA

Extraction protocol

Embryos were washed and dounced in FA buffer (50 mM HEPES/KOH pH 7.5, 1 mM EDTA, 1% Triton X-100, 0.1 % sodium deoxycholate; 150 mM NaCl). 0.1 % sarkosyl was added before sonicating to obtain chromatin fragments of majority between 200-800 bp. 1-2 mg of embryo extract and 3-5 ug of antibody was used per ChIP.
Half of the ChIP DNA and 10-20 ng of input DNA were ligated to Illumina multiplexed adapters and amplified by PCR. Library DNA between 250-500 bp in size was gel purified.

Library strategy

ChIP-Seq

Library source

genomic

Library selection

ChIP

Instrument model

Illumina HiSeq 2000

Data processing

Basecalls were performed using CASAVA version 1.8.
ChIP-seq reads were aligned to the WS220 genome assembly using bowtie version 1.0.1 using default parameters for allowing mismatches in the seed (-n option) and suppressing alignments for reads with more than 4 reportable alignments.
Peaks were called using MACS version 1.4.2 with the following setting: input, genome size (-g ce), format (-f BAM), p-value (-p 1e-10 and -p 1e-5)
Coverage per base was normalized to the genome-wide median coverage (excluding the mitochondrial chromosome). Final ChIP enrichment score per base was obtained by subtracting matching input coverage.
Replicates were merged by averaging coverage at each base position. To determine a set of final peaks per subunits, reads from the replicates were combined using the samtools utility merge version 0.1.19, and MACS was used to call peaks (see above, p-value -p 1e-10). Only those peaks present in the majority of the individual replicates identified below p-value 1e-5 were included in the final peak set.
genome build: WS220
processed data files format and content: Wig files were generated using MACS version 1.4.2, normalized according to the genome-wide mean coverage and the input subtracted. Scores represent the average coverage of each replicate at each base position. Bed files were generated by combining replicate reads and using MACS version 1.4.2 at two different p-values. Only peaks at the more stringent cut-off were inlcuded in the final set that were overlapping with the majority of peaks of each of the replicates at the less stringent p-value.

Submission date

Apr 07, 2015

Last update date

May 15, 2019

Contact name

Sevinc Ercan

Organization name

New York University

Department

Biology

Street address

12 Waverly Place