|
| Status |
Public on Aug 26, 2013 |
| Title |
H3K27me3_ChIPSeq |
| Sample type |
SRA |
| |
|
| Source name |
FBio-CHD5 expressing SH-SY5Y cells
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: neuroblastoma cell line
cell line: SH-SY5Y
expression: FBio-CHD5
treatment: Retinoic Acid treatmeant (8 Days)
chip antibody: H3K27me3 (Millipore, 07-449)
|
| Treatment protocol |
SH-SY5Y cells were treated for 8 days in the presence of 10uM Retinoic Acid (Media was changed on Days 2, 4, 6)
|
| Growth protocol |
SH-SY5Y cells were grown in DMEM supplemented with 10% FBS and 1% Penicillin/Streptomycin
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Differentiated SH-SY5Y cells cells were cross-linked with 1% formaldehyde and chromatin sonicated to an average size of 200-300 bp. Chromatin from FBio-CHD5 and Fbio-Ctrl expressing SH-SY5Y cells was precipitated with magnetic streptavidin beads (Invitrogen) or the H3K27me3 antibody and recovered DNA and an Input DNA control were used for high throughput sequencing.
Libraries were prepared according to Illumina's instructions accompanying the DNA Sample Kit (Part# 0801-0303). Briefly, DNA was end-repaired using a combination of T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenow polymerase) and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (32 to 52 exo minus) and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. After adapter ligation DNA was PCR amplified with Illumina primers for 14 cycles and library fragments of 150-400 bp (insert plus adaptor and PCR primer sequences) were band isolated from an agarose gel. The purified DNA was sent to BGI genomics for sequencing on an Illumina Hi-Seq 2000 platform.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Data processing |
Alignment; 50bp short reads were mapped onto the Human genome (hg19, Feb 2009) using the burrows-wheeler alignment tool, allowing for up to two mismatches in each read.
Peak detection was performed using MACS (Zhang et al. 2008) where the FBio-Ctrl or Input_DNA datasets served as a normalization controls for the FBio-CHD5 and H3K27me3 datasets, respectively.
Tag density files for each dataset were generated using the IGV toolkit.
Genome_build: hg19, Feb 2009
Supplementary_files_format_and_content: tag density files
|
| |
|
| Submission date |
Jun 26, 2013 |
| Last update date |
May 15, 2019 |
| Contact name |
Adrian P. Bracken |
| Organization name |
Trinity College Dublin
|
| Department |
Department of Genetics
|
| Lab |
Bracken Lab
|
| Street address |
1 Lincoln Place
|
| City |
Dublin |
| ZIP/Postal code |
Dublin 2 |
| Country |
Ireland |
| |
|
| Platform ID |
GPL11154 |
| Series (1) |
| GSE48314 |
CHD5 is required for neurogenesis and has a dual role in facilitating gene expression and Polycomb gene repression |
|
| Relations |
| BioSample |
SAMN02213968 |
| SRA |
SRX315189 |
