|
| Status |
Public on Mar 16, 2012 |
| Title |
E10.5_hindlimb_ChIP_input_rep4_index4 |
| Sample type |
SRA |
| |
|
| Source name |
embryonic hindlimb bud, ChIP input
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6J
tissue: hindlimb bud
developmental stage: gestational day E10.5
chip antibody: none
sequencing run type: Paired-end 75 multiplexed
|
| Treatment protocol |
Pregnant mothers were euthanized, and embryos were removed and placed in cold PBS. Forelimb and hindlimb buds were individually removed with forceps.
|
| Growth protocol |
Normal mouse gestation to day 10.5.
|
| 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 procedure included with the Illumina ChIP-Seq kit (IP-102-1001). For multiplexed samples, Illumina Multiplexing adapters and primers (PE-400-1001) were used instead of those provided with the ChIP-Seq kit. Multiplexed samples were paired-end sequenced with an insert size of 300 bp. Standard procedure included with the Illumina mRNA-seq kit (RS-100-0801).
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
Input
|
| Data processing |
ChIP-Seq alignments: ChiP-Seq reads were aligned with bowtie (0.12.3) to the mouse genome (mm9) retaining only uniquely mapped reads (command: bowtie -m 1 -p 8 -B 1 --solexa1.3-quals ~/GENOME/mm9/dna/mm9_nh).
ChIP-Seq peak calling: ChIP peaks were identified by MACS (1.37) against input controls. Default parameters were used for CTCF ChIP-Seq (input control replicates 1 and 2)(command: macs -t ../../s_6_sequence.aligned -c ../../s_5_sequence.aligned --name=Mouse_CTCF_091310_FL_mfold10_p00001 --format=BOWTIE --tsize=74 --gsize=1860000000 --bw=150 -- pvalue=0.00001 --diag --mfold=10 --wig). Nomodel option was selected for H3K27me3 and H3K27ac ChIP seq (input control replicates 3 and 4 respectively)(command: macs -t ../082310/bowtie/s_5_sequence.aligned -c ../082210/bowtie/s_4_sequence.aligned --name=Mouse_h3k27me3_FL1_p00001 --nomodel --shiftsize=1 --format=BOWTIE --tsi ze=74 --gsize=1860000000 --bw=150 --pvalue=0.00001 --diag).
|
| |
|
| Submission date |
Jul 13, 2011 |
| Last update date |
May 15, 2019 |
| Contact name |
Justin Cotney |
| E-mail(s) |
[email protected]
|
| 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 |
GPL13112 |
| Series (1) |
| GSE30641 |
Chromatin state signatures associated with tissue-specific gene expression and enhancer activity in the embryonic limb. |
|
| Relations |
| SRA |
SRX083272 |
| BioSample |
SAMN00668311 |
| Named Annotation |
GSM759873_E10.5_hindlimb_ChIP_input_rep4.bw |
