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Sample GSM1892404

Query DataSets for GSM1892404

Status

Public on Feb 09, 2017

Title

H4K16Ac_ChIPSeq spaced 2

Sample type

SRA

Source name

mushroom body

Organism

Drosophila melanogaster

Characteristics

strain: CS10
chip antibody: anti-H4K16Ac (07-329, Merck Millipore, lot# DAM1780380)
treatment: spaced trained 1d

Treatment protocol

Flies were subjected to spaced trained in which an odor is paired with electrical shocks.

Growth protocol

Flies were raised under a 12 h:12 h, light:dark cycle, at 23℃ and 60% humidity.

Extracted molecule

genomic DNA

Extraction protocol

Heads were collected and homogenized in crosslinking buffer. The nuclei were dissolved in extraction buffer, sonicated to dissociate the individual nuclei, and precipitated with ANTI-FLAG M2 Affinity Gel. The precipitates were rinsed 4 times in extraction buffer, with 5 min nutation at 4℃ between washes, and subjected to ChIP analysis. The purified mushroom body nuclei were sonicated, resulting in fragmentation of DNA ranging from 100 bp to 500 bp. The extracts were centrifuged to remove insoluble materials including the M2 Affinity Gel, and the supernatants were used for immunoprecipitation with specific antibodies.
The ChIP DNA samples prepared were used to prepare a library for SOLiD5500, according to the manufacturer’s instructions.

Library strategy

ChIP-Seq

Library source

genomic

Library selection

ChIP

Instrument model

AB 5500 Genetic Analyzer

Description

H4K16Ac increased sites.txt
all peak H4K16Ac.txt

Data processing

Basecalls and alignment on dm 3 were performed using Lifescope with default configurations
The read with lower mapping quality than 8 was eliminated.
The peaks were called by comparing ChIP DNA with input DNA, using PICS on Strand NGS with a default setting except for the following parameters; for PICS, 120 bp as an average fragment length, 10 bp as a minimum distance between forward and reverse reads, 200 bp as a minimum distance between forward and reverse reads, 100 bp as a window width, with 5% false discovery rate.
The peaks were called by comparing ChIP DNA with input DNA, using ERD on Strand NGS, 2 as an enrichment factor, 100 bp as a window size, 10 bp as a window slide size, 100 bp as a minimum region size.
Overlapping peaks called by PICS and ERD were determined as peaks.
When trained flies and naïve flies were compared, the read counts in the individual peaks were statistically analyzed as follows. First, the centers of the H3K9Ac and H4K16Ac peaks were first extracted and then used to create neighboring 400 bp windows. The mapped reads were counted in these 400 bp windows for each biological replicate of ChIP-seq analysis. The counted read numbers in individual regions were normalized by elav for H3K9Ac ChIP-seq, and gapdh2 for H4K16Ac ChIP. The normalized read numbers in individual windows obtained from the spaced trained flies and the naïve flies were analyzed by Student’s t test (p < 0.05). Standard deviations of the normalized read numbers in individual windows from 3 replicates of naïve flies were 0.07 in H3K9Ac-ChIP-seq and 0.10 in H4K16Ac-ChIP-seq. Significantly increased peaks showing a more than 1.2-fold increase (>2SD) were determined as sites with an increase in H3K9Ac or H4K16Ac.
Genome_build: dm3 (BDGP R5)
Supplementary_files_format_and_content: txt files were generated using Strand NGS; FDR determined by PICS and fold increase determined by ERD were shown. Increase in histone acetylation was determined using three replicates of each naïve and trained flies, and the mean, SD and p value by student t test were shown.
Supplementary_files_format_and_content: all peak H3K9Ac.txt [txt showing all H3K9 peaks from 3 replicates of spaced trained samples (determined by overlaps between PICS and ERD)]
Supplementary_files_format_and_content: all peak H4K16Ac.txt [txt showing all H4K16 peaks from 3 replicates of spaced trained samples (determined by overlaps between PICS and ERD)]
Supplementary_files_format_and_content: H3K9Ac increased sites.txt [txt showing statistically increased sites in H3K9Ac based on three replicates]
Supplementary_files_format_and_content: H4K16Ac increased sites.txt [txt showing statistically increased sites in H4K16Ac based on three replicates]
Supplementary_files_format_and_content: CREB sites.txt [txt showing CREB binding sites (determined by overlaps between PICS and ERD)]
Supplementary_files_format_and_content: CRTC sites.txt [txt showing CRTC binding sites (determined by overlaps between PICS and ERD)]

Submission date

Sep 24, 2015

Last update date

May 15, 2019

Contact name

Yukinori Hirano

E-mail(s)

yukinori@ust.hk

Organization name

The Hong Kong University of Science and Technology

Department

LIFS