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Sample GSM442430 Query DataSets for GSM442430
Status Public on Aug 22, 2009
Title E0-4_CBP_3_of_3_rep_2
Sample type genomic
 
Channel 1
Source name E0-4_CBP
Organism Drosophila melanogaster
Characteristics development stage: E0-4
antibody: CBP
antibody provider: Dr. Mattias Mannervik in Stockholm University
test: ChIP
Growth protocol 1. y; bw cn sp flies are cultivated in cages with apple juice agar plates covered with yeast powder. 2. For a 0-4 hours egg laying, new plates are added after a 2hours pre-egglaying at 10 am and flies are allowed to lay eggs for 4 hours. 3. Embryos are collected at 1.45 pm using a filter mesh and a brush. They are rinsed with EWB and cross-linked at 2 pm in 1.8% formaldehyde at room temperature for 15 min.
Extracted molecule genomic DNA
Extraction protocol Drosophila samples are crushed in presence of 1.8% of formaldehyde for 15 minutes. After washing, the material is sonicated for 15 minutes to solubilize the chromatin. After centrifugation, the supernatant containing soluble chromatin is collected for IPs. Specific antibodies are added to the chromatin extract while one aliquot will be treated for DNA extraction corresponding to the reference samples. ChIP are carried overnight at room temperature. Protein A Sepharose beads are used to pull down the chromatin bound antibodies. After washes and finally elution, the DNA corresponding to the IP'd material is extracted and purified. Amplification of the DNA is done by linker-mediated PCR and used for labeling.
Label Cy3
Label protocol 2 ug of amplified IP DNA are used for random primer labeling reaction (invitrogen). Cy3-dCTP is incorporated for the ChIP samples while Cy5-dCTP is incorporated for the Input chromatin sample. After washing of the labeled material on Centricon columns, 5ug of labeled DNA is used for hybridization.
 
Channel 2
Source name E0-4_CBP
Organism Drosophila melanogaster
Characteristics reference: Input
development stage: E0-4
Growth protocol 1. y; bw cn sp flies are cultivated in cages with apple juice agar plates covered with yeast powder. 2. For a 0-4 hours egg laying, new plates are added after a 2hours pre-egglaying at 10 am and flies are allowed to lay eggs for 4 hours. 3. Embryos are collected at 1.45 pm using a filter mesh and a brush. They are rinsed with EWB and cross-linked at 2 pm in 1.8% formaldehyde at room temperature for 15 min.
Extracted molecule genomic DNA
Extraction protocol Drosophila samples are crushed in presence of 1.8% of formaldehyde for 15 minutes. After washing, the material is sonicated for 15 minutes to solubilize the chromatin. After centrifugation, the supernatant containing soluble chromatin is collected for IPs. Specific antibodies are added to the chromatin extract while one aliquot will be treated for DNA extraction corresponding to the reference samples. ChIP are carried overnight at room temperature. Protein A Sepharose beads are used to pull down the chromatin bound antibodies. After washes and finally elution, the DNA corresponding to the IP'd material is extracted and purified. Amplification of the DNA is done by linker-mediated PCR and used for labeling.
Label Cy5
Label protocol 2 ug of amplified IP DNA are used for random primer labeling reaction (invitrogen). Cy3-dCTP is incorporated for the ChIP samples while Cy5-dCTP is incorporated for the Input chromatin sample. After washing of the labeled material on Centricon columns, 5ug of labeled DNA is used for hybridization.
 
 
Hybridization protocol The hybridization solution is composed of 5 ug each of Cy3- and Cy5-labeled DNA, 50 ug of salmon sperm DNA, Agilent Blocking Agent and Agilent Hyb Buffer. The mixture is heated to 95*C for 3 min and then 37*C for 30 minutes immediately prior to hybridization. Samples are hybridized for 40 hours at 65*C. After hybridization the slides are washed in Agilent Wash Buffer 1 for 5 minutes at room temperature and then in Agilent Wash Buffer 2 for 5 minutes at 31*C.
Scan protocol Channel 1 & 2 are for the Agilent arrays are scanned on Agilent's DNA Microarray Scanner G2505B at a resolution of 5 microns. Data is extracted from the images using Agilent's Feature Extraction Software 9.5.3.1.
Description E0-4_CBP
Data processing We preprocessed microarray data to normalize it and reduce experimental noise as described in Qi et al. (Qi et al., Nature biotechnology 2006). In short, the raw intensities from each channel was divided by the median intensity from that channel before computing a ratio to arrive at a median adjusted ratio. We further subtracted average value of negative control probes on the array and then added one as a regularization parameter.
 
Submission date Aug 19, 2009
Last update date Aug 19, 2013
Contact name Kevin P. White
E-mail(s) kpwhite@uchicago.edu
Organization name University of Chicago
Department Institute for Genomics and Systems Biology
Street address 900 E. 57th STR. 10th FL.
City Chicago
State/province IL
ZIP/Postal code 60615
Country USA
 
Platform ID GPL6951
Series (2)
GSE15292 Drosophila at different time points of development: ChIP-chip, ChIP-seq, RNA-seq
GSE15427 ChIP-chip of CBP/p300 in Drosophila at different time points of development

Data table header descriptions
ID_REF
VALUE Normalized log ratios between the Cy3 channel and the Cy5 channel intensities.

Data table
ID_REF VALUE
125813 -0.206317313359464
211284 -0.326082176598275
116584 0.217311174659948
3459 0.486553295239785
204749 -0.413288497780823
36909 0.0414530464841802
219799 -0.399343489692732
115177 -0.248500848064969
191452 0.0743385646831636
80970 0.239309356969766
70521 0.462714320640323
173479 -0.181517072703527
114558 0.382951167628762
224367 -0.435233996433036
30370 -0.14912626072872
11193 0.246419880544657
55621 0.81636376649956
23522 0.311967744625818
135143 0.467527942394002
241647 -0.290728842881521

Total number of rows: 237869

Table truncated, full table size 5852 Kbytes.




Supplementary file Size Download File type/resource
GSM442430.txt.gz 66.6 Mb (ftp)(http) TXT
Processed data included within Sample table

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