|
| Status |
Public on Jul 02, 2013 |
| Title |
human_E33_rep2_input |
| Sample type |
SRA |
| |
|
| Source name |
human_E33
|
| Organism |
Homo sapiens |
| Characteristics |
developmental stage: gestational day E33
tissue: forelimb and hindlimb
|
| Treatment protocol |
Human samples were obtained from the Human Developmental Biological Resource (HDBR).
|
| Growth protocol |
Normal Gestatation
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
For each ChIP-Seq, limb bud tissue was crosslinked with 1% formaldehyde at room temperature and stored at -80°C. Chromatin was extracted and sheared by sonication. Soluble chromatin was combined with Protein G Dynabeads prebound with appropriate antibodies at 4C overnight. Beads were collected with magnet and washed 5x. Chromatin was eluted with TE+1%SDS at 65C for 10 minutes. Crosslinks were reversed at 65C overnight, and then chromatin was purified with the PCR cleanup kit (Qiagen).
Standard Paired End Illumina Multiplexing protocols
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
processed data file (Anonymized alignments): E33_rep2_input_300_uniq.bed
|
| Data processing |
Basecalls with CASAVA-1.8.2
ChIP-seq and RNA-Seq reads were aligned to the hg19, rheMac2, or mm9 genome assembly using Bowtie (0.12.7) with the -m1 flag.
Aligned ChIP-Seq reads in each sample were scanned along the genome in 500bp windows at 25bp step-size. For each window, the total number of reads from H3K27ac and control experiment were counted respectively. Raw read number in control experiment was then scaled to match the sequencing depth of H3K27ac experiment. The number of expected reads in a window based on uniform distribution of total mapped reads along a specific chromosome was also calculated. Then the significance of enrichment was calculated using a Poisson model where the null model is the larger number of control or expected read counts in the window. Significantly enriched windows (p-value≤10-5) within 1kb of each other were then merged.
Signal plots were generated by extending reads to 300bp and counting number of reads at each position in wiggle format. Wiggle tracks were then converted to bigwig using wigToBigWig from the Kent utilities.
Genome_build: hg19, rheMac2, or mm9
Supplementary_files_format_and_content: No raw data are provided for human samples. Human alignments were anonymized by removing sequence information and converting to bed format
Supplementary_files_format_and_content: Enriched Region files are in bed format.
Supplementary_files_format_and_content: Bigwig files were generated with wigToBigWig with the -clip flag.
|
| |
|
| Submission date |
Nov 20, 2012 |
| Last update date |
Jul 02, 2013 |
| Contact name |
Justin Cotney |
| E-mail(s) |
cotney@uchc.edu
|
| Organization name |
UConn Health
|
| Department |
Genetics and Genome Sciences
|
| Lab |
Cotney Lab
|
| Street address |
400 Farmington Ave.
|
| City |
Farmington |
| State/province |
CT |
| ZIP/Postal code |
06030-6403 |
| Country |
USA |
| |
|
| Platform ID |
GPL11154 |
| Series (1) |
| GSE42413 |
The Evolution of Lineage-Specific Regulatory Activities in the Human Embryonic Limb |
|
| Relations |
| BioSample |
SAMN02197220 |
