|
| Status |
Public on Nov 09, 2011 |
| Title |
EWS502_FAIRE |
| Sample type |
SRA |
| |
|
| Source name |
Ewing Sarcoma cells
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: EWS502
chip antibody: none
|
| Treatment protocol |
A short hairpin region complementary to the 3' untranslated region of FLI1 together with PCR-generated HA-EWS-FLI and HA-FLI1 were cloned into pLL5.5 (Rubinson et al. 2003). Lentivirus was produced in HEK293T cells as described (Rubinson et al. 2003). EWS502 or HUVEC cells were infected with lentivirus in the presence of polybrene (6 µg/mL) for 3 h after which media was changed. Chromatin or RNA was isolated at 72 h.
|
| Growth protocol |
EWS502 were cultured in RPMI 1640 supplemented with 15% FBS, HUVEC cells were cultured in Vasculife Basal Media (Lifeline Technologies) supplemented with 10% FBS. Both cell lines were maintained at standard growth conditions of 37o and 5% CO2
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
For chromatin immunoprecipitation, chromatin was isolated and immunoprecipitation was performed as described (Davis et al. 2006) using 2 ug of anti-HA antibody (Abcam ab9110), anti-H3K4me1 (Abcam ab8895), anti-H3K4me2 (Abcam ab32356), anti-H3K4me3 (Abcam ab12209), or anti-H3K27me3 (Millipore 07-449). FAIRE was performed on three independent cell harvests as previously described (Giresi et al. 2007). All libraries were prepared as per Illumina's recommendations, including two rounds of purification with Agencourt AMPure XP beads.
|
| |
|
| Library strategy |
FAIRE-seq |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina Genome Analyzer IIx |
| |
|
| Description |
Formaldehyde-Assisted Isolation of Regulatory Elements
|
| Data processing |
Reads from chromatin immunoprecipitations were aligned to the reference human genome (hg18) with Bowtie using default parameters, and unambiguously placed reads were retained. Biological replicates were then merged and reads were extended in silico to a final length of 200 bp. Any extended reads that overlapped large-scale repetitive elements were then removed. Reads from FAIRE were allowed to potentially map to up to four genomic locations, but the best scoring alignment was chosen. Biological replicates were then merged and reads were extended in silico to a final length of 134 bp. Any extended reads that overlapped large-scale repetitive elements were then removed.
|
| |
|
| Submission date |
Sep 02, 2011 |
| Last update date |
May 12, 2023 |
| Contact name |
Ian Jonathan Davis |
| E-mail(s) |
ian_davis@med.unc.edu
|
| Phone |
919-966-5360
|
| Organization name |
University of North Carolina
|
| Department |
Genetics, Pediatrics
|
| Street address |
450 West Drive
|
| City |
Chapel Hill |
| State/province |
NC |
| ZIP/Postal code |
27599 |
| Country |
USA |
| |
|
| Platform ID |
GPL10999 |
| Series (1) |
| GSE31838 |
Tumor-specific retargeting of an oncogenic transcription factor chimera results in dysregulation of chromatin and transcription |
|
| Relations |
| SRA |
SRX096369 |
| SRA |
SRX096370 |
| BioSample |
SAMN00716145 |
