|
| Status |
Public on Feb 11, 2010 |
| Title |
IMR90hTert_Input |
| Sample type |
SRA |
| |
|
| Source name |
IMR90hTert_Input
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: IMR90hTert
antibody: None
|
| Treatment protocol |
MR90HTert cells were cross-linked for seven minutes in 1% formaldehyde. Following cell and nuclear lysis as described, extracted chromatin was sonicated to an average size of 200-600 base pairs and used for immunoprecipitations. The sonicated chromatin was mixed with various antibodies and incubated on a rotating wheel overnight at 4˚C. Immunoprecipitated material was recovered by addition of 10 µl of protein A agarose beads (pre-blocked with 10µg/ml of BSA and salmon sperm DNA) and incubation for 1 h at room temperature on a rotating wheel. The beads were then washed with dialysis and wash buffer {O'Geen, 2006 #64}. De-crosslinking, RNase A and proteinase K treatments, and DNA purification were conducted as described {O'Geen, 2006 #64}.
|
| Growth protocol |
The chromatin immunoprecipitations (ChIPs) were performed with exponentially growing IMR90HTert cells, which are diploid fibroblast-like cells established from fetal female lung tissue and transfected with the gene coding for human Telomerase reverse Transcriptase.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
At least 10 ng was used to prepare sequencing libraries with the Illumina ChIP-Seq Sample Preparation Kit according to the protocol supplied with the reagents. Aliquots of the sequencing libraries were then loaded into Genome Analyzer flow cells and sequenced with the Genome Analyzer II (Illumina) using the 36 Cycle Sequencing Kit v2 (Catalog number FC-204-2036). As controls, aliquots of the input DNA (collected after the crosslinking and sonication steps) were also subjected to ultra high throughput sequencing.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina Genome Analyzer II |
| |
|
| Description |
High-throughput sequencing of control input DNA (collected after the crosslinking and sonication steps)
|
| Data processing |
Alignment : The sequenced tags were mapped onto the unmasked genome build hg18 using the fetchGWI software and allowing 1000 repeats on the genome. The tags mapping more than once onto the genome were then attributed a weight corresponding to the number of times the tag was sequenced divided by the number of genomic matches . The files with these weighted repeated tags as well as the unique tags were then used for peak detection
Peak detection : peaks were determined with the sissrs software (www.rajajothi.com/sissrs/). Peaks common to the control input material and the ChIP material, as well as peaks mapping in satellite and micro-satellite repeats and peaks mapping in 18 and 28S rRNA sequences, were eliminated from the analysis. This first analysis resulted in the identification of peaks. The obtained peaks were then quantified using the protocol described in the paper describing this dataset
|
| |
|
| Submission date |
Sep 21, 2009 |
| Last update date |
May 15, 2019 |
| Contact name |
Nicolo Riggi |
| Organization name |
CHUV
|
| Department |
Département de Pathologie Expérimentale
|
| Lab |
Institut universitaire de pathologie
|
| Street address |
Bugnon 25
|
| City |
Lausanne |
| State/province |
VD |
| ZIP/Postal code |
1011 |
| Country |
Switzerland |
| |
|
| Platform ID |
GPL9115 |
| Series (1) |
| GSE18184 |
Genome-wide localization of the RNA polymerase III transcription machinery in human cells |
|
| Relations |
| SRA |
SRX016558 |
| BioSample |
SAMN00008676 |
