|
| Status |
Public on Dec 31, 2016 |
| Title |
HOXD11_ChIP_Replicate1 |
| Sample type |
SRA |
| |
|
| Source name |
Chicken Micromass primary cells
|
| Organism |
Gallus gallus |
| Characteristics |
cell type: MSCs derived from embryonal HH25 limb bud
infection: 3xFLAG-HOXD11
days in culture: 6 days
antibody: FLAG (Sigma Mouse monoclonal M2; F3165)
|
| Treatment protocol |
Cells were infected with RCASBP viruses carrying different transgenes.
|
| Growth protocol |
MSCs were isolated from the HH24 chicken limb buds, infected with an RCASBP(A) or (B) virus carrying the transgene on interest. Afterwards cells were cultured for 6 days with the DMEM:HAM F11 media (supplemented with 10% Fetal Bovine Serum, 0,2% Chicken Serum, 1% Penicilin-Streptomicin and 1% L-glutamine).
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Nuclei were isolated with three different lysis buffers as in Lee et al., 2006 and sonicated in the lysis buffer 3 until the chromatin was disrupted to the size between 200 and 300bp.
Libraries were prepared using NEBNext ChIP-seq Library Prep Master Mix for Illumina (New England Biolabs), fragments between 300-450bp were selected and sequenced on a Illumina HiSeq 1500 using Illumina TruSeq v3 chemistry.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 1500 |
| |
|
| Description |
HOXD11.bw
HOXD11_peaks.bed
HOXD11-CTCF_peaks.bed
|
| Data processing |
Reads were filtered by quality using FASTX Toolkit 0.0.13 (-q 28 -p 50 -v -o X -Q 33)
Samples were aligned to the Galgal4 genome using BWA 0.5.9 for Illumina (-n 0 -n 0.04 -o 1 -e -1 -d 16 -i 5 -l-1 -k 2 -M 3 -O 11 -E 4)
Lines were selected with the following pattern: XT:A:U|^@SQ|^@PG
sam files were converted to bam with the samtools version 0.1.18
bam files were merged with Picard 1.56 (beta) version and peaks were called on the pooled bam files using MACS2 (2.0.10.20120913) with the p-value cut off at 1e-1, tag size 50 and the rest of the default options. Afterwards the peaks were ranked according to the p-value and only the reproducible peaks from the list were seleced. Finally the peaks were filtered to discard the peaks located on mitochondrial or random chromosomes. IDR pipeline was used to determine the number of reproducible peaks for every sample which was done as in: Kundaje et al., 2012 ( https://sites.google.com/site/anshulkundaje/projects/idr )
Genome_build: Galgal4
Supplementary_files_format_and_content: bed files are produced as described above. Shared peaks are considered shared only is the summits are at the most 200bp away or closer. Bigwig files are normalised reads per milion and are extended 150bp. Bigwig files were generated using the samtools and bedtools.
Supplementary_files_format_and_content: CTCF_peaks.bed: HOX-CTCF co-bound peaks
|
| |
|
| Submission date |
Aug 26, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Daniel Murad Ibrahim |
| E-mail(s) |
ibrahim@molgen.mpg.de, daniel.ibrahim@bih-charite.de
|
| Phone |
030 84131516
|
| Organization name |
Berlin Institute of Health
|
| Department |
Center for Regenerative Therapies
|
| Street address |
Augustenburger Platz 1
|
| City |
Berlin |
| State/province |
Berlin |
| ZIP/Postal code |
13353 |
| Country |
Germany |
| |
|
| Platform ID |
GPL21476 |
| Series (2) |
| GSE86088 |
Genome-wide binding of posterior HOXA/D transcription factors reveals subgrouping and association with CTCF [ChIP-Seq] |
| GSE86089 |
Genome-wide binding of posterior HOXA/D transcription factors reveals subgrouping and association with CTCF |
|
| Relations |
| BioSample |
SAMN05710581 |
| SRA |
SRX2055089 |
