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| Status |
Public on Apr 14, 2021 |
| Title |
H3K9me3_veh rep1 |
| Sample type |
SRA |
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| Source name |
H3K9me3_veh rep1
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| Organism |
Homo sapiens |
| Characteristics |
cell type: LNCap cell line
antibody: Anti-H3K9me3
vendor: Millipore
catalog #: 17-10242
treatment: DMSO
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| Treatment protocol |
10 nM DHT was added before cell collection for MNase-seq/ MNase-ChIP-seq of histone markers assay.
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| Growth protocol |
LNCaP cells were cultured in 10% FBS RPMI 1640 medium, 1:2.5 split every 2 days. Cells are cultured in 5% charcoal stripped FBS RPMI-1640 medium
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
extract 1: For ChIP-exo, after treatment with 10 nm DHT or vehicle for 4 h, LNCaP cells were crosslinked with 1% formaldehyde for 10 min. Sonicated chromatin was enriched by immunoprecipatation with an anti-GATA2 antibody and T4 DNA polymerase, T4 PNK, and Klenow DNA Polymerase were used together for end polishing. The ligation step was performed with less reducing agent. Protein A Dynal magnetic beads were washed using modified RIPA buffer (50mM Tris-HCL pH 7.8, 1mM EDTA, 0.25% Na Deoxycholate, 1% NP-40, 0.5M LiCl). The library was amplified with only 10 or 12 PCR cycles, and prepared without gel-based size selection.
extract2: For Mnase-ChIP-seq, chromatin was isolated from freshly cultured cells and Mono-nucleosomes with solubilized chromatin was achieved by MNase digestion at 37 °C, then immunoprecipitated with antibody-conjugated magnetic beads. DNA is phenol extracted and ethanol precipitated.
Libraries were prepared according to Illumina's instructions accompanying the DNA Sample Kit (Part#15023092). Briefly, DNA was end-repaired, phosphorylated ends were treated with Klenow fragment and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. After adapter ligation, the DNA was size selected from an agarose gel and was PCR amplified with Illumina primers for 15 cycles. Libraries were sequenced on the Hi-seq 3000 following the manufacturer's protocols.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 3000 |
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| Description |
Extraction protocol 2
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| Data processing |
hIP-seq reads were aligned to the hg19 genome assembly using Bowtie2 version with parameters '-k 2 -v 2' to report up to two valid alignments
Read with multiple valid alignments were removed using a Perl program
Uniquely mapped reads from replicates of the same sample were pooled
Peaks were called using MACS2 with parameters '-m 3,30 -g hs -p 1e-8'
ChIP-exo data were processed using ePEST with parameters '-p 1e-8 -R 25 -c 0.05 -k 2.0'
BigWig file was generated by deeptools for visualization
Genome_build: GRCh37/hg19
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| Submission date |
Apr 20, 2020 |
| Last update date |
Mar 28, 2022 |
| Contact name |
Tianbao Li |
| E-mail(s) |
tianbaoli89@gmail.com
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| Organization name |
University of Texas Health Science Center at San Anotnio
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| Department |
Molecular Medicine
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| Lab |
Victor Jin
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| Street address |
7703 Floyd Curl
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| City |
San Antonio |
| State/province |
Texas |
| ZIP/Postal code |
78229 |
| Country |
USA |
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| Platform ID |
GPL21290 |
| Series (1) |
| GSE148935 |
LNCap cell nucleosome footprint elicits novel noncanonical GATA2 pioneer model |
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| Relations |
| BioSample |
SAMN14644724 |
| SRA |
SRX8142295 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4486249_H3K9me3_veh_rep1.bigWig |
193.4 Mb |
(ftp)(http) |
BIGWIG |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
| Processed data provided as supplementary file |
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