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Status |
Public on Feb 09, 2017 |
Title |
CREB_ChIPSeq naïve 3 |
Sample type |
SRA |
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Source name |
mushroom body
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Organism |
Drosophila melanogaster |
Characteristics |
strain: CS10 chip antibody: anti-CREB (lab made antibodies) treatment: naïve
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Treatment protocol |
Flies were subjected to spaced trained in which an odor is paired with electrical shocks.
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Growth protocol |
Flies were raised under a 12 h:12 h, light:dark cycle, at 23℃ and 60% humidity.
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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.
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Library strategy |
ChIP-Seq |
Library source |
genomic |
Library selection |
ChIP |
Instrument model |
Illumina MiSeq |
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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)
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Submission date |
May 16, 2016 |
Last update date |
May 15, 2019 |
Contact name |
Kunio Ihara |
E-mail(s) |
ihara@gene.nagoya-u.ac.jp
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Phone |
81527894532
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Organization name |
Nagoya University
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Department |
Center for Gene Research
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Street address |
Furo-cho 1, Chikusa-ku
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City |
Nagoya |
State/province |
Aichi |
ZIP/Postal code |
4648602 |
Country |
Japan |
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Platform ID |
GPL16479 |
Series (1) |
GSE81456 |
Genome-wide maps of CREB/CRTC binding in Drosophila mushroom body. |
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Relations |
BioSample |
SAMN05001337 |
SRA |
SRX1763407 |
Supplementary data files not provided |
SRA Run Selector |
Processed data are available on Series record |
Raw data are available in SRA |
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