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Sample GSM386219 Query DataSets for GSM386219
Status Public on Mar 28, 2009
Title E-4-8h_H3K27AC_2_of_3_rep_1
Sample type genomic
 
Channel 1
Source name ChIP, H3K27AC in Drosophila embryos at 4-8 hours of development, E-4-8h_H3K27AC_2_of_3_rep_1
Organism Drosophila melanogaster
Characteristics test: ChIP
antibody: H3K27Ac
antibody manufacturer: Abcam
antibody catalog number: ab4729
antibody lot number: 34629
tissue: embryos
time point: 4-8 hours of development
replicate: 2_of_3_rep_1
Treatment protocol No treatment
Growth protocol 1. y; bw cn sp flies are cultivated in cages with apple juice agar plates covered with yeast powder. 2. For a 4-8 hours egg laying, new plates are added after a 2hours pre-egglaying at 9 am and flies are allowed to lay eggs for 4 hours. 3. Embryos are collected at 4.45 pm using a filter mesh and a brush. They are rinsed with EWB and cross-linked at 5 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 Input, H3K27AC in Drosophila embryos at 4-8 hours of development, E-4-8h_H3K27AC_2_of_3_rep_1
Organism Drosophila melanogaster
Characteristics reference: Input
tissue: embryos
time point: 4-8 hours of development
replicate: 2_of_3_rep_1
Treatment protocol No treatment
Growth protocol 1. y; bw cn sp flies are cultivated in cages with apple juice agar plates covered with yeast powder. 2. For a 4-8 hours egg laying, new plates are added after a 2hours pre-egglaying at 9 am and flies are allowed to lay eggs for 4 hours. 3. Embryos are collected at 4.45 pm using a filter mesh and a brush. They are rinsed with EWB and cross-linked at 5 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 ChIP-chip of H3K27AC in Drosophila embryos at 4-8 hours of development, E-4-8h_H3K27AC_2_of_3_rep_1
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 Mar 25, 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 GPL6950
Series (2)
GSE15292 Drosophila at different time points of development: ChIP-chip, ChIP-seq, RNA-seq
GSE15430 ChIP-chip of H3K27Ac 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
187694 -0.588785536847071
234359 0.104208706202799
25818 -0.284974117511947
41947 -0.055514326357362
205970 -0.316864940758866
130688 0.282418088923387
69684 0.390802451729449
171093 -0.100079326086638
101858 3.59786192357142
216572 0.657781164168843
190489 -0.504251486030444
44898 -0.062832136177715
24398 4.88164187326709
116207 -0.218937177824465
137069 -0.41404776760797
100722 -0.231790107331842
184904 0.362524441636843
100206 1.20241637074529
138224 -0.403766855675049
207216 -0.142998820568561

Total number of rows: 237894

Table truncated, full table size 5793 Kbytes.




Supplementary file Size Download File type/resource
GSM386219.txt.gz 23.5 Mb (ftp)(http) TXT
Raw data provided as supplementary file
Processed data included within Sample table

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