|
| Status |
Public on Jan 31, 2015 |
| Title |
LSD2-KD H3K4me1 |
| Sample type |
SRA |
| |
|
| Source name |
HepG2, LSD2-KD H3K4me1
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: HepG2
variation: LSD2-knockdown
chip antibody: anti-H3K4me1 (abcam, ab8895)
|
| Treatment protocol |
For the knockdown experiments, specific siRNAs were introduced to the cells using RNAiMAX reagent (Invitrogen) when they were approximately 50% confluent.
|
| Growth protocol |
HepG2 cells were cultured in high glucose (25mM D-glucose) Dulbecco-modified eagle’s medium (DMEM) (Sigma) supplemented with 10% (v/v) heat-inactivated fetal bovine serum and penicillin/streptomycin.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Crosslinked nuclear extracts were subjected to DNA fragmentation by Covaris S220 sonifier, and chromatin-DNA complexes were pulled down using specific antibodies.
Libraries were prepared using an Ion Plus Fragment Kit (Lifetechnologies, Part no. 4471252) according to the manufacturer's instructions. Briefly, ChIP DNA was end-repaired using an end repair enzyme, and was purified with two rounds of AMPure XP bead capture to size‐select fragments approximately 100–250 bp in length. After adapter ligation, DNA was PCR amplified with Ion primers for 13 cycles. The purified DNA was checked for quality on Agilent 2100 Bioanalyzer instrument with an Agilent High Sensitivity DNA kit. The library DNA was approximately 170–220 bp in length. Libraries ware quantified using quantitative real-time PCR with the Ion Library Quantitation Kit (Part no. 4468802). Diluted librarie DNA was subjected to emulsion PCR using Ion OneTouch System and sequenced on the Ion PGM and Ion Proton semiconductor sequencers (Lifetechnologies) according to the manufacturer's protocols.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Ion Torrent Proton |
| |
|
| Data processing |
Basecalls performed using Torrent Suite (Life technologies)
All sequence data obtained from Ion Proton were merged before alignment onto the human reference genome hg19 using the BWA algorithm (CLC Genomics Workbench Software).
Duplicate reads and low read/mapping quality reads were trimmed out with the CLC Genomics Workbench Software.
Peak detection was done using the MACS algorithm in Avadis NGS software. H3K4me1 binding sites in LSD2-depleted HepG2 cells were detected based on the H3K4me1 peaks significantly enriched over H3K4me1 peaks in control HepG2 cells at a cutoff value of p=10^-5.
Genome_build: hg19
Supplementary_files_format_and_content: BED file generated using MACS algorithm in Avadis NGS software.
|
| |
|
| Submission date |
Jul 28, 2014 |
| Last update date |
May 15, 2019 |
| Contact name |
Shinjiro Hino |
| E-mail(s) |
s-hino@kumamoto-u.ac.jp
|
| Organization name |
Kumamoto University
|
| Lab |
Medical Cell Biology
|
| Street address |
2-2-1 Honjo
|
| City |
Kumamoto |
| ZIP/Postal code |
860-0811 |
| Country |
Japan |
| |
|
| Platform ID |
GPL17303 |
| Series (2) |
| GSE59695 |
Role of histone lysine demethylase LSD2 in hepatic metabolism |
| GSE59830 |
ChIP-seq analysis of LSD2/H3K4me1 interaction in control and LSD2-knockdowned HepG2 cells |
|
| Relations |
| BioSample |
SAMN02943106 |
| SRA |
SRX663303 |
