|
| Status |
Public on Sep 19, 2011 |
| Title |
H9 anti Myc |
| Sample type |
genomic |
| |
|
| Channel 1 |
| Source name |
ChIP DNA H9 anti Myc Santa Cruz
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: human embryonic cell line
antibody: Anti Myc Santa Cruz (Cat # SC-764)
|
| Growth protocol |
All cells were grown in defined growth media and were in log phase collected for ChIP
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Cells were treated with 1% formaldehyde to crosslink DNA and proteins. Cells were washed and nuclei were isolated, lysed and chromatin was sheared. Chromatin was immunoprecipitated with either anti MYC or anti rabbit IgG. Purified DNA was amplified and labeled following Affymetrix Chromatin Immunoprecipitatin Assay protocol.
|
| Label |
biotin
|
| Label protocol |
7.5 ug of amplified ChIP DNA was fragmented using UDG and APE at 37C for 60 minutes, and terminal labeled with biotinylated nucleotide and terminal DNA transferase at 37C for 60 minutes.
|
| |
|
| Channel 2 |
| Source name |
ChIP DNA H9 anti rabbit IgG
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: Human H9 embryonic cell line
antibody: Anti rabbit IgG (Cat#SC-2027)
|
| Growth protocol |
All cells were grown in defined growth media and were in log phase collected for ChIP
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Cells were treated with 1% formaldehyde to crosslink DNA and proteins. Cells were washed and nuclei were isolated, lysed and chromatin was sheared. Chromatin was immunoprecipitated with either anti MYC or anti rabbit IgG. Purified DNA was amplified and labeled following Affymetrix Chromatin Immunoprecipitatin Assay protocol.
|
| Label |
biotin
|
| Label protocol |
7.5 ug of amplified ChIP DNA was fragmented using UDG and APE at 37C for 60 minutes, and terminal labeled with biotinylated nucleotide and terminal DNA transferase at 37C for 60 minutes.
|
| |
|
| |
| Hybridization protocol |
The labeled DNA was hybridized to the Affymetrix GeneChip human promoter arrays for 16hr at 45 C with constant rotation (60 rpm). Affymetrix Fluidics Station 450 was used to wash and stain the Chips, removing the non-hybridized target and incubating with a streptavidin-phycoerythrin conjugate to stain the biotinilated cDNA. The staining was further amplified using goat IgG as blocking reagent and biotinilated anti-streptavidin antibody (goat), followed by a second staining step with a streptavidin-phycoerythrin conjugate.
|
| Scan protocol |
Fluorescence was detected using the Affymetrix G3000 GeneArray Scanner and image analysis of each GeneChip was perfomed through the Affymetrix GeneChip Command Console version 2.0 (AGCC v2.0) software from Affymetrix, using the standard default settings
|
| Description |
control CEL file KZel-MycChIP-H9aHGF-3a-HuProm1.CEL was also used and is provided on the Series record as supplementary files.
Myc ChIP with Santa Cruz antibody in H9 human ES cells, Rep 1-3, promoter
|
| Data processing |
Data was quantile normalized and analyzed with TileMapv2 implemented in CisGenome software. Log2(IP/Input) fold changes (fc) and TileMap moving average (ma) statistics were reported in BAR files.
fc.bar and ma.bar are generated by CisGenome software. fc.bar contains log2(IP/Control) fold changes for each probe. ma.bar contains a moving average statistic (i.e. the MA statistic in TileMap) for each probe. bed files contain all high quality binding sites reported in the paper.
|
| |
|
| Submission date |
Sep 07, 2011 |
| Last update date |
Sep 19, 2011 |
| Contact name |
Hongkai Ji |
| E-mail(s) |
hji@jhsph.edu
|
| Organization name |
Johns Hopkins Bloomberg School of Public Health
|
| Department |
Department of Biostatistics
|
| Street address |
615 North Wolfe Street, Rm E3638
|
| City |
Baltimore |
| State/province |
MD |
| ZIP/Postal code |
21205 |
| Country |
USA |
| |
|
| Platform ID |
GPL5082 |
| Series (2) |
| GSE31964 |
MYC binding in human B cells (P493-6) and ESC (H9) |
| GSE32220 |
Cell-type independent MYC target genes reveal a primordial signature involved in biomass accumulation |
|
