|
| Status |
Public on Dec 04, 2009 |
| Title |
pol II unstimulated |
| Sample type |
genomic |
| |
|
| Channel 1 |
| Source name |
pol II ChIP DNA from unstimulated T cell
|
| Organism |
Homo sapiens |
| Characteristics |
antibody: pol II
cell line: Jurkat T cells
treatment: unstimulated
|
| Treatment protocol |
Jurkat cells were unstimulated
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
1x10^7 cells were fixed with 1% formaldehyde in culture medium for 5 min at room temperature followed by quenching with 0.125 M glycine for 15 min. The cells were washed twice with ice-cold PBS and the ChIP pellets are harvested and stored at -80 C before performing the ChIP assay. The detailed ChIP assays are published in Byun et al PNAS 2009.
|
| Label |
Cy5
|
| Label protocol |
ChIP DNA was directly labeled by Klenow (New England Biolabs) random priming with Cy5 per manufacturer's protocol (http://www.nimblegen.com/products/lit/lit.html).
|
| |
|
| Channel 2 |
| Source name |
input DNA
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: Jurkat T cells
treatment: unstimulated
|
| Treatment protocol |
Jurkat cells were unstimulated
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
1x10^7 cells were fixed with 1% formaldehyde in culture medium for 5 min at room temperature followed by quenching with 0.125 M glycine for 15 min. The cells were washed twice with ice-cold PBS and the ChIP pellets are harvested and stored at -80 C before performing the ChIP assay. The detailed ChIP assays are published in Byun et al PNAS 2009.
|
| Label |
Cy3
|
| Label protocol |
ChIP DNA was directly labeled by Klenow (New England Biolabs) random priming with Cy5 per manufacturer's protocol (http://www.nimblegen.com/products/lit/lit.html).
|
| |
|
| |
| Hybridization protocol |
The labeled ChIP DNA was precipitated with 0.1 volume 5M NaCl and 1 volume isopropanol, and hybridized in 45 ul of buffer containing 20% formamide, 1.2 M betaine, 0.1 ug/ul herring sperm DNA and 10 ug of human COT1 DNA (Invitrogen). Arrays were hybridized in Maui hybridization stations for 16-18 h at 42C, and then washed in 42C 0.2% SDS/0.2x SSC, room temperature 0.2x SSC, and 0.05x SSC. Hybridization buffers and washes were completed using manufacturer's protocols (http://www.nimblegen.com/products/lit/lit.html)
|
| Scan protocol |
Arrays were scanned on an Axon 4000B scanner per manufacturer's protocol (http://www.nimblegen.com/products/lit/lit.html).
|
| Description |
none
|
| Data processing |
Arrays were processed using Nimblegen's standard protocol for Nimblescan 2.4 ChIP data extraction.
GFF files include scaled, log2 (ChIP/Input) ratio in the 6th column. Positional information is based on human genome build 35.
|
| |
|
| Submission date |
Dec 03, 2009 |
| Last update date |
Dec 03, 2009 |
| Contact name |
Jung S Byun |
| E-mail(s) |
byunjs@mail.nih.gov
|
| Phone |
3014352890
|
| Organization name |
NCI/NIH
|
| Lab |
Genetics Branch
|
| Street address |
37 convent drive, Bldg 37, Rm 1042
|
| City |
Bethesda |
| State/province |
MD |
| ZIP/Postal code |
20892 |
| Country |
USA |
| |
|
| Platform ID |
GPL3497 |
| Series (1) |
| GSE19311 |
Chip-chip from Jurkat with p300 and pol II |
|
