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Sample GSM2152730 Query DataSets for GSM2152730
Status Public on Feb 09, 2017
Title CREB_ChIPSeq spaced 2
Sample type SRA
 
Source name mushroom body
Organism Drosophila melanogaster
Characteristics strain: CS10
chip antibody: anti-CREB (lab made antibodies)
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 Miseq, according to the manufacturer’s instructions.
 
Library strategy ChIP-Seq
Library source genomic
Library selection ChIP
Instrument model Illumina MiSeq
 
Data processing Basecalls and alignment on dm 3 were performed using CLCbio 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 were obtained in the 1 kb window covering entire genome and analyzed by DESeq2, to determine the region enriched with CREB/CRTC in a specific group of samples. The region with increased or decreased CREB/CRTC binding were determined if the region were defined by the CREB/CRTC binding sites in the above criteria.
Genome_build: dm3
Supplementary_files_format_and_content: txt files were generated using Strand NGS and DESeq2; score and FDR determined by PICS, and adjusted p value and fold change determined by DESeq2
Supplementary_files_format_and_content: CREB CRTC binding analyzed by DESeq2.txt: txt showing statistically alteration in CREB/CRTC binding on three replicates
Supplementary_files_format_and_content: CREB sites.txt: txt showing CREB binding sites (determined by peak calling at least in 2 samples out of 3 biological replicates)
Supplementary_files_format_and_content: CRTC sites.txt: txt showing CRTC binding sites (determined by peak calling at least in 2 samples out of 3 biological replicates)
 
Submission date May 16, 2016
Last update date May 15, 2019
Contact name Kunio Ihara
E-mail(s) ihara@gene.nagoya-u.ac.jp
Phone 81527894532
Organization name Nagoya University
Department Center for Gene Research
Street address Furo-cho 1, Chikusa-ku
City Nagoya
State/province Aichi
ZIP/Postal code 4648602
Country Japan
 
Platform ID GPL16479
Series (1)
GSE81456 Genome-wide maps of CREB/CRTC binding in Drosophila mushroom body.
Relations
BioSample SAMN05001339
SRA SRX1763409

Supplementary data files not provided
SRA Run SelectorHelp
Processed data are available on Series record
Raw data are available in SRA

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