|
| Status |
Public on May 23, 2007 |
| Title |
Embryonic Fibroblast Line (MEF2) H3me3K4 Promoter Methylation ChIP-Chip (2/2) |
| Sample type |
genomic |
| |
|
| Channel 1 |
| Source name |
Embryonic Fibroblast (MEF2)
|
| Organism |
Mus musculus |
| Characteristics |
Elute from a histone H3 trimehtylated K4 chromatin immunoprecipitation
|
| Growth protocol |
Standard ESC media conditions
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Genome wide chromatin analysis ChIP was performed with about 1 million cells following the protocol on www.upstate.com. The cells were crosslinked with formaldehyde for 10 min at room temperature, subsequently lysed in 10mM Tris-EDTA pH 8.0 with 1%SDS, and sonicated on ice 6 times at 15 second pulses interrupted by 45 second pauses. Clarified sheared chromatin was immunoprecipitated with antibodies to H3me3K4 (Abcam 8580) overnight at 4 C, collected with protein A beads for 2 hours, washed twice for 5 min and eluted with buffers (recipes on the Upstate website). Eluates were reverse crosslinked, RNAse and proteinase K treated, and DNA was purified using the Qiagen PCR purification kit.
|
| Label |
Cy5
|
| Label protocol |
10ng of each immuoprecipitated sample and corresponding inputs were amplified using the Whole Genome Amplification Kit (Sigma), and 2ug of amplified material was labeled with Cy5 (Perkin Elmer) using the Bioprime Kit (Invitrogen).
|
| |
|
| Channel 2 |
| Source name |
Embryonic Fibroblast (MEF2)
|
| Organism |
Mus musculus |
| Characteristics |
Input for histone H3 trimehtylated K4 chromatin immunoprecipitation
|
| Growth protocol |
Standard ESC media conditions
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Genome wide chromatin analysis ChIP was performed with about 1 million cells following the protocol on www.upstate.com. The cells were crosslinked with formaldehyde for 10 min at room temperature, subsequently lysed in 10mM Tris-EDTA pH 8.0 with 1%SDS, and sonicated on ice 6 times at 15 second pulses interrupted by 45 second pauses.
|
| Label |
Cy3
|
| Label protocol |
10ng of each immuoprecipitated sample and corresponding inputs were amplified using the Whole Genome Amplification Kit (Sigma), and 2ug of amplified material was labeled with Cy3 (Perkin Elmer) using the Bioprime Kit (Invitrogen).
|
| |
|
| |
| Hybridization protocol |
Hybridization onto the mouse promoter array, which includes 244K features distributed onto two slides (Agilent –G4490), washing and scanning were carried out according to the manufacturers instructions.
|
| Scan protocol |
According to Agilent Scanning Software
|
| Description |
Embryonic Fibroblast Line (MEF2) H3me3K4 Promoter Methylation ChIP-Chip (2/2)
|
| Data processing |
Log ratios were calculated by the Agilent Feature Extraction software and automatically underwent Lowess normalization.
|
| |
|
| Submission date |
May 15, 2007 |
| Last update date |
May 23, 2007 |
| Contact name |
Kathrin Plath |
| Organization name |
UCLA
|
| Department |
Biological Chemistry
|
| Lab |
BSRB 390B
|
| Street address |
615 Charles E. Young Drive South
|
| City |
Los Angeles |
| State/province |
CA |
| ZIP/Postal code |
90095-1737 |
| Country |
USA |
| |
|
| Platform ID |
GPL4129 |
| Series (1) |
| GSE7815 |
Histone H3K4 and H3K27 trimethylation analysis in ES cells, Mefs and induced pluriptent cells. |
|
