|
| Status |
Public on May 12, 2009 |
| Title |
H3K4me3_E0-12h |
| Sample type |
genomic |
| |
|
| Channel 1 |
| Source name |
H3K4me3_E0-12h
|
| Organism |
Drosophila melanogaster |
| Characteristics |
antibody: H3K4me3
antibody manufacturer: Abcam
antibody catalog number: ab8580
antibody lot number: 411277
tissue: Embryos
time point: 0-12h
|
| 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 0-12 hours egg laying, new plates are added after a 2hours pre-egglaying at 9 pm and flies are allowed to lay eggs for 12 hours. 3. Embryos are collected at 8.45 am using a filter mesh and a brush. They are rinsed with EWB and cross-linked at 9 am 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 IPed material is extracted and purified. Amplification of the DNA is done by linker-mediated PCR and used for labeling.
|
| Label |
biotin
|
| Label protocol |
Affymetrix Labeling Protocol (TdT labeling)
|
| |
|
| Channel 2 |
| Source name |
input
|
| Organism |
Drosophila melanogaster |
| Characteristics |
reference: input
|
| 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 0-12 hours egg laying, new plates are added after a 2hours pre-egglaying at 9 pm and flies are allowed to lay eggs for 12 hours. 3. Embryos are collected at 8.45 am using a filter mesh and a brush. They are rinsed with EWB and cross-linked at 9 am 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 IPed material is extracted and purified. Amplification of the DNA is done by linker-mediated PCR and used for labeling.
|
| Label |
biotin
|
| Label protocol |
Affymetrix Labeling Protocol (TdT labeling)
|
| |
|
| |
| Hybridization protocol |
Affymetrix Hybridization Protocol - Hybridizations were performed at the Functional Genomics Facility (FGF) at the University of Chicago.
|
| Scan protocol |
Affymetrix Protocol - Scans were performed at the Functional Genomics Facility (FGF) at the University of Chicago.
|
| Description |
H3K4me3_E0-12h
test: A2_H3K4me3.CEL
test: C2_H3K4Me31.CEL
test: C2_H3K4Me32.CEL
reference: A2_Mock.CEL
reference: C2_Mock1.CEL
reference: C2_Mock2.CEL
|
| Data processing |
.CEL files generated by the scanner are used as Input files in the MAT software to generate the signal files that are used by CisGenome to identify peaks.
.bed and .bar were generated with MAT.
|
| |
|
| Submission date |
May 06, 2009 |
| Last update date |
Feb 02, 2015 |
| Contact name |
Kevin P. White |
| E-mail(s) |
[email protected]
|
| 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 |
GPL6629 |
| Series (1) |
| GSE15980 |
ChIP-chip of H3K4me3 in Drosophila embryos at E0-12h time point. |
|
| Relations |
| Named Annotation |
GSM400362_MAT_H3K4me3_E0-12h.bed.gz |
