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| Status |
Public on Jan 24, 2017 |
| Title |
BAT_adipocyte_H3K27ac_400K |
| Sample type |
SRA |
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| Source name |
BAT
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| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6J
tissue: adipose
chip antibody: H3K27ac (Active Motif, 39133)
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| Growth protocol |
Mice were maintained on a standard chow diet (8664 Harlan Teklad, 6.4% w/w fat) under a regular 12h light/12h dark cycle at constant temperature (23°C).
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Tissues were dounce homogenized, and cross-linked nuclei were isolated and sorted by BD FACS Aria II using mCherry fluorescence. Sorted nuclei were sheared by Covaris E220 and used for ChIP for overnight. Immunoprecipitates were washed and subjected to elution and reverse crosslinking. DNA was then extracted by AMPure XP beads according to the manufacturer’s manual.
Extracted DNA (1-10ng, or all if less) was used to generate sequencing libraries by following the “on-bead” sequencing library preparation method. Briefly, DNA was processed through end repair/phosphorylation using the End-It DNA End-Repair Kit (Epicentre), A-tailing using the Klenow Fragment (NEB M0212) and index adaptor ligation using the Quick Ligase (NEB M2200). AMPure XP beads were left in all the reactions to clean up the DNA using PEG(Polyethylene Glycol 8000)/NaCl solution. After ligation, DNA was eluted from AMPure XP beads and then PCR-amplified using the PfuUltra II Hotstart PCR Master Mix (Agilent 600850). Gel electrophoresis/extraction was performed using the E-Gel EX Agarose Gels (Invitrogen) and MinElute Gel Extraction (Qiagen) to select library fragments between 250 and 600bp. Quantity and quality of the libraries were analyzed by Qubit and Agilent Bioanalyzer, respectively, and the libraries were pooled at a final concentration of 12pM and sequenced by HiSeq2500 or NextSeq 500 systems.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NextSeq 500 |
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| Data processing |
ChIP-seq data were aligned using Bowtie2 to the mm9 mouse genome
Duplicates and low quality reads were removed by Picard (http://picard.sourceforge.net)
Reads were processed using samtools (Li et al., 2009)
Peaks were called by MACS2 (Zhang et al., 2008), and low peaks (4<fold higher coverage than whole cell extract (WCE)) were removed.
Reads were assigned to peaks, normalized and quantified using featureCounts and EdgeR. Low signal peaks (log2 CPM<2) were removed, and differential peaks were defined at log2 FC≥1 and log2 FDR≤0.25.
For ChIP-seq track visualization, reads were processed to the BigWig file format using bedtools (Quinlan and Hall, 2010) and bedGraphToBigWig (Kent et al., 2010).
Processed Bigwig files were viewed in WashU Epigenome Browser (Zhou and Wang, 2012).
Genome_build: mm9
Supplementary_files_format_and_content: BigWig files
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| Submission date |
Dec 19, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Evan Rosen |
| E-mail(s) |
erosen@bidmc.harvard.edu
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| Organization name |
Beth Israel Deconess Medical Center
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| Department |
Endocrinology
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| Lab |
Rosen Lab
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| Street address |
3 Blackfan Cir
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| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02215 |
| Country |
USA |
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| Platform ID |
GPL19057 |
| Series (1) |
| GSE92590 |
Simultaneous transcriptional and epigenomic profiling from specific cell types within heterogeneous tissues in vivo |
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| Relations |
| BioSample |
SAMN06166476 |
| SRA |
SRX2436358 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM2433310_BAT_adipocyte_H3K27ac_400K.bw |
188.0 Mb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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