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Sample GSM2120825

Query DataSets for GSM2120825

Status

Public on Nov 14, 2016

Title

testis_input_DNA_R2

Sample type

SRA

Source name

Adult testis, input

Organism

Danio rerio

Characteristics

strain/background: AB
genotype/variation: wild type
Sex: male
age: adult
tissue: testis
chip antibody: none

Extracted molecule

genomic DNA

Extraction protocol

Whole brains, hearts, intestines and testis were dissected from adult male AB zebrafish (brains, hearts and testis = 10-month-old; hearts and intestines = 1-year-old). Two biological replicates were prepared from brain, heart (1-year-old), intestine and testis, whereas one biological replicate was prepared from 10-month-old heart. Each biological replicate was prepared using: 12 brains, 20 hearts, 5 intestines and 8 testis. All tissues were homogenized and cross-linked in 1% formaldehyde, washed and lysed. Chromatin was sheared using a Covaris S220 ultrasonicator to a DNA fragment size of 175 bp (10-month-old samples) or 200 bp (1-year-old samples). ChIP-seq was performed as previously described (Guenther et al. 2008) using 5 ug Abcam H3K27ac antibody (ab4729, lot# GR259887-1) bound to Dynal Protein A linked beads (Invitrogen). Reverse cross-linked and phenol:chloroform purified chromatin was used for single-end library preparation following standard Illumina protocols.
DNA fragments with an average size of 175 bp or 200 bp were used for single-end library preparation following standard Illumina protocols.

Library strategy

ChIP-Seq

Library source

genomic

Library selection

ChIP

Instrument model

Illumina HiSeq 2500

Description

Processed data file: adult_testis_H3K27ac_drerio_super_enhancers.bed
Processed data file: adult_testis_H3K27ac_drerio_typical_enhancers.bed
Processed data file: adult_testis_drerio_H3K27ac_normalized.wig

Data processing

Basecalls were performed using CASAVA.
ChIP-seq reads were aligned to the Zv9 genome assembly using Bowtie2-2.1.0 configured for global alignment and allowing 1 mismatch in the seed alignment (seed size = 22). If a read aligned more than once in the genome, only the alignment with the best score was saved for further analyses. Aligned reads were saved in BAM format using samtools (version 0.1.8).
BAM files from biological replicates were merged with samtools and converted to BED format with bedtools (v2.24.0).
Peak calling was performed using SICER (V1.1) with the following parameters: redundancy threshold = 1, window size = 200, fragment size = 175 or 200, effective genome fraction = 0.7, gap size = 600 and FDR = 0.05.
After peak calling, peaks were discarded if 50% of the peak overlapped with the regions comprising 2 Kb upstream and 2 Kb downstream of TSSs (based on RefSeq annotations) and if the peak summit was within these regions.
To identify typical enhancers and super-enhancers, the ROSE algorithm was applied (Whyte et al. 2013; Lovén et al. 2013), setting the -t parameter to 2000.
Genome_build: Zv9
Supplementary_files_format_and_content: BED files with annotations for typical enhancers and super-enhancers were generated using ROSE. Scores represent the H3K27ac density in each enhancer region.
Supplementary_files_format_and_content: WIG file with the H3K27ac profile generated by SICER and normalized by library size per million.

Submission date

Apr 12, 2016

Last update date

May 15, 2019

Contact name

Yuvia Alheli Perez Rico

E-mail(s)

yuvia.perez-rico@embl.de

Organization name

Institut Curie

Street address

26 rue d'Ulm