|
| Status |
Public on Aug 20, 2017 |
| Title |
ChIP-seq_H3K27me3_Col_rep2 |
| Sample type |
SRA |
| |
|
| Source name |
Seedlings, WT, H3K27me3 ChIP
|
| Organisms |
Arabidopsis thaliana; Homo sapiens |
| Characteristics |
ecotype: Col
genotype/variation: wildtype
tissue: seedlings
age: 10 days
human cell line: HEK293
chip antibody: H3K27me3 (Millipore 07-449)
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Mononucleosomes from 10-day-old Arabidopsis seedlings and human HEK293 cells generated by micrococcal nuclease (MNase) digestion were subjected to ChIP. To generate Arabidopsis mononucleosomes, nuclei were extracted from 10-day-old seedlings, and were subsequently subjected to MNase digestion in N buffer (15mM Tris pH7.5, 250mM sucrose, 60mM KCl, 15mM NaCl, 5mM MgCl2, 1mM CaCl2, 1mM DTT, 10mM ß-mercaptoethanol, protease inhibitor cocktail). EDTA and EGTA were added to a final concentration of 10mM each to stop the MNase digestion. Nuclei were lysed by adding NaCl to a final concentration of 500mM, and soluble materials containing mononucleosomes were recovered after centrifugation. To generate mononucleosomes from human HEK293 cells, nuclei were extracted by hypotonic lysis and treated with MNase in N buffer. DNA was extracted from a portion of mononucleosomes to determine the concentration of mononucleosomes. Same amount of mononucleosomes from different Arabidopsis samples were mixed with a constant amount of human mononucleosomes before immunoprecipitation with anti-H3K27me3 antibody.
Sequencing libraries were prepared using a TruSeq DNA sample prep kit (Illumina).
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Description |
Human DNA was used to calculate the reference derived normalization factor.
|
| Data processing |
Reads were mapped to the Arabidopsis genome (TAIR10) or human genome (hg38/GRCh38) using Bowtie2.
The number of H3K27me3 reads aligned to human genome were first normalized based on the number of input reads, and then was used to calculate reference derived normalization factor.
BedGraph files were created using the deepTools utility bamCoverage with a bin size of 10bp.
The number of reads in each bin in the bedGraphs were then scaled by the reference derived normalization factor to generate reference-adjusted reads per million (RRPM).
H3K27me3 enriched regions were identified using MACS2.
Genome_build: TAIR10 (Arabidopsis), hg38/GRCh38 (human)
Supplementary_files_format_and_content: *bigwig: Normalized coverage of H3K27me3.
|
| |
|
| Submission date |
Jul 11, 2017 |
| Last update date |
Feb 06, 2020 |
| Contact name |
Danhua Jiang |
| E-mail(s) |
dhjiang@genetics.ac.cn
|
| Organization name |
Institute of Genetics and Developmental Biology
|
| Street address |
No.1 West Beichen Road
|
| City |
Beijing |
| ZIP/Postal code |
100101 |
| Country |
China |
| |
|
| Platform ID |
GPL23696 |
| Series (2) |
| GSE101220 |
DNA replication-coupled histone modification maintains Polycomb gene silencing in plants [ChIP-Rx] |
| GSE101221 |
DNA replication-coupled histone modification maintains Polycomb gene silencing in plants |
|
| Relations |
| Reanalyzed by |
GSE144900 |
| BioSample |
SAMN07343966 |
| SRA |
SRX2996432 |
