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| Status |
Public on May 31, 2023 |
| Title |
ECh_Abcg2N_K4me3_2_052721 |
| Sample type |
SRA |
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| Source name |
Heart
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| Organism |
Mus musculus |
| Characteristics |
tissue: Heart
cell type: Endothelial Cells
genotype: Abcg2CreErt;ROSATdTomato
treatment: 50mg/kg tamoxifen injection
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| Treatment protocol |
50mg/kg tamoxifen i.p injected 1 day before TdTomato+CD31+CD45-Ter119- AbcVESC (Abcg2p) and TdTomato-CD31+CD45-Ter119- mature endothelial cells (Abcg2n) were flow sorted from Abcg2TT mice heart
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| Growth protocol |
Tissue samples from 8 week old ABCG2TT mice
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Chromatin immunoprecipitation was performed with ULI-NChIP protocol. 10,000 FACS isolated ECs were used per reaction. Cells were treated with the nuclei extraction buffer (10 mM Tris-HCl pH8.5, 140 mM NaCl, 5 mM MgCl2, 0.6% NP40, 1x protease inhibitor cocktail) and chromatin was fragmented for 10 min using MNase at 25°C and diluted in NChIP immunoprecipitation buffer (10 mM Tris-HCl pH8.0, 2 mM EDTA, 90 mM NaCl, 0.1% Triton X-100, 0.1% DOC, 1x protease inhibitor cocktail). Chromatin was incubated with 1μg of antibody bound to 11μl of protein A Dynabeads (Thermo Fisher Scientific, 10002D) and rotated overnight at 4°C. Then it was washed twice with 200 μl low-salt wash buffer (20 mM Tris-HCl pH8.0, 2mM EDTA, 150mM NaCl, 1% Triton X-100, 0.1% SDS), and twice with 200μl high-salt wash buffer (20mM Tris-HCl pH8.0, 2mM EDTA, 500mM NaCl, 1% Triton X-100, 0.1% SDS). DNA was eluted from chromatin in 100μl hot elution buffer (100mM NaHCO3, 1% SDS) for 2 hr at 65°C, then purified by phenol:chloroform and precipitated by isopropanol overnight at -20°C. The DNA was resuspended in 10mM Tris-HCl pH 8.5. ChIP was performed using the following antibodies: anti-H3K4me3 (Cell signaling Technology, 9727), anti- H3K27me3 (Millipore, 07-449).
ChIP sequence libraries were generated using the KAPA Hyper Prep Kit
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NovaSeq 6000 |
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| Description |
Abcg2- EC K4me3 2
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| Data processing |
Sequencing service was performed on an Illumina Hiseq 4000 sequencer
ChIP-seq reads were aligned to the Mus musculus genome (mm9, NCBI Build 37) using the BWA program (version 0.5.9), and PCR duplicates were removed by Picard (version 1.69) (http://picard.sourceforge.net/).
Unique reads that mapped to a single best-matching location with no more than 4% of the read length of mismatches were kept and used to study genome-wide enrichment of specific histone modification.
Sequence data was visualized with IGV by normalizing to 1 million reads
The SICER (version 1.1) algorithm was applied to the ChIP-seq data, with sequencing data from input DNA as control to identify genomic enrichment (peak) of specific histone modifications (FDR<0.01). To identify differentially enriched regions, SICER-df was used with FDR<0.01.
Promoters were defined as fragments between upstream 2 kb and transcription start site (TSS). The ChIP-seq read counts within promoters were quantified by HOMER (version 4.10.4)
Assembly: mm9
Supplementary files format and content: bigWig
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| Submission date |
Feb 23, 2023 |
| Last update date |
May 31, 2023 |
| Contact name |
Yang Lin |
| Organization name |
Weill Cornell Medicine
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| Street address |
455 Main Street 06W
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| City |
New York |
| State/province |
NY |
| ZIP/Postal code |
10044 |
| Country |
USA |
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| Platform ID |
GPL24247 |
| Series (1) |
| GSE225987 |
Abcg2 expressing VESC in adult heart |
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| Relations |
| BioSample |
SAMN33426842 |
| SRA |
SRX19488328 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM7061347_ECh_Abcg2N_K4me3_2_052721.bw |
14.1 Mb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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