|
| Status |
Public on Mar 16, 2012 |
| Title |
E11.5_forelimb_H3K27ac_ChIP_seq_index6_mate |
| Sample type |
SRA |
| |
|
| Source name |
embryonic forelimb bud
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6J
tissue: forelimb bud
developmental stage: gestational day E11.5
method: ChIP-seq
chip antibody: anti-H3K27ac
antibody vendor: Abcam
antibody catalog #: ab4729
antibody lot#: GR46123
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. Total RNA was extracted with RNEasy kit (Qiagen) for RNA-seq. 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 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 then chromatin was purified with PCR cleanup kit (Qiagen).
|
| Growth protocol |
Normal mouse gestation to day 10.5.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Standard procedure included with 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 Illumina mRNA-seq kit (RS-100-0801).
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
Chromatin IP against H3K27ac
|
| 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 |
Feb 14, 2012 |
| 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 |
SRX120060 |
| BioSample |
SAMN00790463 |
| Named Annotation |
GSM875385_E11.5_forelimb_H3K27ac_ChIP_seq.bw |
| Named Annotation |
GSM875385_E11.5_forelimb_H3K27ac_ChIP_seq_peaks.bed.gz |
